Macrocycles useful in the treatment of Alzheimer&#39;s disease

ABSTRACT

The present invention are macrocycles of the formula (IX): 
                 
 
for treating Alzheimer&#39;s disease and other similar diseases. These compounds include inhibitors of the beta-secretase enzyme that are useful in the treatment of Alzheimer&#39;s disease and other diseases characterized by deposition of A beta peptide in a mammal. The compounds of the invention are useful in pharmaceutical compositions and methods of treatment to reduce A beta peptide formation.

This application claims the benefit of U.S. Provisional PatentApplication Nos. 60/297,546 filed Jun. 12, 2001, and 60/333,083 filedNov. 19, 2001.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The invention relates to substituted cyclic amides and such compoundsthat are useful for the treatment of Alzheimer's disease. Morespecifically, the invention relates to such compounds that are capableof inhibiting beta-secretase, an enzyme that cleaves amyloid precursorprotein to produce amyloid beta peptide (A beta), a major component ofthe amyloid plaques found in the brains of Alzheimer's sufferers.

2. Description of the Related Art

Alzheimer's disease (AD) is a progressive degenerative disease of thebrain primarily associated with aging. Clinical presentation of AD ischaracterized by loss of memory, cognition, reasoning, judgment, andorientation. As the disease progresses, motor, sensory, and linguisticabilities are also affected until there is global impairment of multiplecognitive functions. These cognitive losses occur gradually, buttypically lead to severe impairment and eventual death in the range offour to twelve years.

Alzheimer's disease is characterized by two major pathologicobservations in the brain: neurofibrillary tangles and beta amyloid (orneuritic) plaques, comprised predominantly of an aggregate of a peptidefragment know as A beta. Individuals with AD exhibit characteristicbeta-amyloid deposits in the brain (beta amyloid plaques) and incerebral blood vessels (beta amyloid angiopathy) as well asneurofibrillary tangles. Neurofibrillary tangles occur not only inAlzheimer's disease but also in other dementia-inducing disorders. Onautopsy, large numbers of these lesions are generally found in areas ofthe human brain important for memory and cognition.

Smaller numbers of these lesions in a more restricted anatomicaldistribution are found in the brains of most aged humans who do not haveclinical AD. Amyloidogenic plaques and vascular amyloid angiopathy alsocharacterize the brains of individuals with Trisomy 21 (Down'sSyndrome), Hereditary Cerebral Hemorrhage with Amyloidosis of theDutch-Type (HCHWA-D), and other neurodegenerative disorders.Beta-amyloid is a defining feature of AD, now believed to be a causativeprecursor or factor in the development of disease. Deposition of A betain areas of the brain responsible for cognitive activities is a majorfactor in the development of AD. Beta-amyloid plaques are predominantlycomposed of amyloid beta peptide (A beta, also sometimes designatedbetaA4). A beta peptide is derived by proteolysis of the amyloidprecursor protein (APP) and is comprised of 39-42 amino acids. Severalproteases called secretases are involved in the processing of APP.

Cleavage of APP at the N-terminus of the A beta peptide bybeta-secretase and at the C-terminus by one or more gamma-secretasesconstitutes the beta-amyloidogenic pathway, i.e. the pathway by which Abeta is formed. Cleavage of APP by alpha-secretase produces alpha-sAPP,a secreted form of APP that does not result in beta-amyloid plaqueformation. This alternate pathway precludes the formation of A betapeptide. A description of the proteolytic processing fragments of APP isfound, for example, in U.S. Pat. Nos. 5,441,870; 5,721,130; and5,942,400.

An aspartyl protease has been identified as the enzyme responsible forprocessing of APP at the beta-secretase cleavage site. Thebeta-secretase enzyme has been disclosed using varied nomenclature,including BACE, Asp, and Memapsin. See, for example, Sindha et al.,1999, Nature 402:537-554 (p501) and published PCT applicationWO00/17369.

Several lines of evidence indicate that progressive cerebral depositionof beta-amyloid peptide (A beta) plays a seminal role in thepathogenesis of AD and can precede cognitive symptoms by years ordecades. See, for example, Selkoe, 1991, Neuron 6:487. Release of A betafrom neuronal cells grown in culture and the presence of A beta incerebrospinal fluid (CSF) of both normal individuals and AD patients hasbeen demonstrated. See, for example, Seubert et al., 1992, Nature359:325-327.

It has been proposed that A beta peptide accumulates as a result of APPprocessing by beta-secretase, thus inhibition of this enzyme's activityis desirable for the treatment of AD. In vivo processing of APP at thebeta-secretase cleavage site is thought to be a rate-limiting step in Abeta production, and is thus a therapeutic target for the treatment ofAD. See for example, Sabbagh, M., et al., 1997, Alz. Dis. Rev. 3, 1-19.

BACE1 knockout mice fail to produce A beta, and present a normalphenotype. When crossed with transgenic mice that over express APP, theprogeny show reduced amounts of A beta in brain extracts as comparedwith control animals (Luo et al., 2001 Nature Neuroscience 4:231-232).This evidence further supports the proposal that inhibition ofbeta-secretase activity and reduction of A beta in the brain provides atherapeutic method for the treatment of AD and other beta amyloiddisorders.

At present there are no effective treatments for halting, preventing, orreversing the progression of Alzheimer's disease. Therefore, there is anurgent need for pharmaceutical agents capable of slowing the progressionof Alzheimer's disease and/or preventing it in the first place.

Compounds that are effective inhibitors of beta-secretase, that inhibitbeta-secretase-mediated cleavage of APP, that are effective inhibitorsof A beta production, and/or are effective to reduce amyloid betadeposits or plaques, are needed for the treatment and prevention ofdisease characterized by amyloid beta deposits or plaques, such as AD.

SUMMARY OF THE INVENTION

The invention provides compounds of formula (IX):

wherein

-   U is-   - - - is an optional bond;-   J is —CH₂OH or —NH—R_(c) when - - - is not a bond, or absent    when - - - is a bond;-   G is OH when - - - is not a bond or —O— when - - - is a bond;-   R is hydrogen or C₁-C₆ alkyl;-   B represents    -   —(CR₄R₅)_(m)—; or    -   C₂-C₆ alkenyl optionally substituted with one, two or three        groups independently selected from R₆, R_(6′) and R_(6″); or    -    where        -   q is 0 or 1; and        -   the “e” ring is            -   aryl or heteroaryl, each of which is optionally                substituted with one, two or three groups independently                selected from R₆, R_(6′), and R_(6″); or            -   a carbocyclic ring having three, four, five or six atoms                in which one, two or three of such atoms are optionally                hetero atoms independently selected from O, N, and S and                where the carbocyclic ring is optionally substituted                with one, two or three groups independently selected                from R₆, R_(6′) and R_(6″);-   m is 1-6;-   R₄ and R₅ are independently H, C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆    alkynyl, C₁-C₆ haloalkyl, C₃-C₇ cycloalkyl, C₄-C₁₂ cycloalkylalkyl,    C₁-C₆ alkoxyalkyl, or C₃-C₆ cycloalkyl;-   X represents    -   —(CR₄R₅)_(m)—; or    -   C₃-C₆ alkenyl optionally substituted with one, two or three        groups independently selected from R₆, R_(6′) and R_(6″); or    -   —CH₂C(═O)NHCHR_(a)—; or    -    where        -   q is 0 or 1; and        -   the “g” ring is a carbocyclic ring having three, four, five            or six atoms in which one, two or three of such atoms are            optionally hetero atoms independently selected from O, N,            and S and where the carbocyclic ring is optionally            substituted with one, two or three groups independently            selected from R₆, R_(6′) and R_(6″);-   R_(a) is a D or L amino acid side chain;-   Y is hydrogen, C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, C₁-C₆    haloalkyl, C₃-C₇ cycloalkyl, C₄-C₁₂ cycloalkylalkyl, C₁-C₆    alkoxyalkyl, C₃-C₆ cycloalkyl, or-   Y together with the carbon to which it is attached is a D or L amino    acid side chain;-   R₆, R_(6′) and R_(6″) independently are    -   C₁-C₆ alkyl optionally substituted with one, two or three groups        independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,        —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino; or    -   C₂-C₆ alkenyl or C₂-C₆ alkynyl, each of which is optionally        substituted with one, two or three groups independently selected        from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,        amino, and mono- or dialkylamino; or    -   —(CH₂)₀₋₄—O—(C₁-C₆ alkyl), where the alkyl portion is optionally        substituted with one, two, three, four, or five groups        independently selected from halogen; or    -   —OH, —NO₂, halogen, —CO₂H, —C≡N, —(CH₂)₀₋₄—CO—NR₈R₉,        —(CH₂)₀₋₄—CO—(C₁-C₁₂ alkyl), —(CH₂)₀₋₄—CO—(C₂-C₁₂ alkenyl),        —(CH₂)₀₋₄—CO—(C₂-C₁₂ alkynyl), —(CH₂)₀₋₄—CO—(C₃-C₇ cycloalkyl),        —(CH₂)₀₋₄—R_(aryl), —(CH₂)₀₋₄—R_(heteroaryl),        —(CH₂)₀₋₄—R_(heterocyclyl), —(CH₂)₀₋₄—CO—R_(aryl),        —(CH₂)₀₋₄—CO—R_(heteroaryl), —(CH₂)₀₋₄—CO—R_(heterocyclyl),        —(CH₂)₀₋₄—CO—R₁₀, —(CH₂)₀₋₄—CO—O—R₁₁, —(CH₂)₀₋₄—SO₂—NR₈R₉,        —(CH₂)₀₋₄—SO—(C₁-C₈ alkyl), —(CH₂)₀₋₄—SO₂₋(C₁-C₁₂ alkyl),        —(CH₂)₀₋₄—SO₂—(C₃-C₇ cycloalkyl), —(CH₂)₀₋₄—N(H or        R₁₁)—CO—O—R₁₁, —(CH₂)₀₋₄—N(H or R₁₁)—CO—N(R₁₁)₂,        —(CH₂)₀₋₄—N—CS—N(R₁₁)₂, —(CH₂)₀₋₄—N(—H or R₁₁)—CO—R₈,        —(CH₂)₀₋₄—NR₈R₉, —(CH₂)₀₋₄—R₁₀, —(CH₂)₀₋₄—O—CO—(C₁-C₆ alkyl),        —(CH₂)₀₋₄—)—P(O)—(O—R_(aryl))₂, —(CH₂)₀₋₄—O—CO—N (R₁₁)₂,        —(CH₂)₀₋₄—O—CS—N(R₁₁)₂, —(CH₂)₀₋₄—O—(R₁₁),        —(CH₂)₀₋₄—O—(R₁₁)—COOH, —(CH₂)₀₋₄—S—(R₁₁), C₃-C₇ cycloalkyl,        —(CH₂)₀₋₄—N(—H or R₁₁)—SO₂—R₇, or —(CH₂)₀₋₄—C₃-C₇ cycloalkyl;-   R₈ and R₉ are the same or different and represent —H, —C₃-C₇    cycloalkyl, —(C₁-C₂ alkyl)-(C₃-C₇ cycloalkyl), —(C₁-C₆    alkyl)-O—(C₁-C₃ alkyl), —C₁-C₆ alkenyl, —C₁-C₆ alkynyl, or —C₁-C₆    alkyl chain with one double bond and one triple bond; or    -   —C₁-C₆ alkyl optionally substituted with —OH or —NH₂; or    -   —C₁-C₆ alkyl optionally substituted with one, two or three        groups independently selected from halogen; or    -   heterocyclyl optionally substituted with one, two or three        groups selected from halogen, amino, mono- or dialkylamino, —OH,        —C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆ alkyl, —SO₂—N(C₁-C₆ alkyl)₂,        —SO₂—(C₁-C₄ alkyl), —CO—NH₂, —CO—NH—C₁-C₆ alkyl, oxo,        —CO—N(C₁-C₆ alkyl)₂,        -   C₁-C₆ alkyl optionally substituted with one, two or three            groups independently selected from C₁-C₃ alkyl, halogen,            —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or            dialkylamino,        -   C₂-C₆ alkenyl or C₂-C₆ alkynyl, each of which is optionally            substituted with one, two or three groups independently            selected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃,            C₁-C₃ alkoxy, amino, and mono- or dialkylamino, and        -   C₁-C₆ alkoxy optionally substituted with one, two or three            groups independently selected from halogen; or    -   aryl or heteroaryl, each of which is optionally substituted with        one, two or three groups independently selected from halogen,        amino, mono- or dialkylamino, —OH, —C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆        alkyl, —SO₂—N(C₁-C₆ alkyl)₂, —SO₂—(C₁-C₄ alkyl), —CO—NH₂,        —CO—NH—C₁-C₆ alkyl, and —CO—N(C₁-C₆ alkyl)₂,        -   C₁-C₆ alkyl optionally substituted with one, two or three            groups independently selected from C₁-C₃ alkyl, halogen,            —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or            dialkylamino,        -   C₂-C₆ alkenyl or C₂-C₆ alkynyl, each of which is optionally            substituted with one, two or three groups independently            selected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃,            C₁-C₃ alkoxy, amino, and mono- or dialkylamino, and        -   C₁-C₆ alkoxy optionally substituted with one, two or three            of halogen;-   R₁₀ is heterocyclyl optionally substituted with one, two, three or    four groups independently selected from C₁-C₆ alkyl;-   R₁₁ is C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, C₃-C₇ cycloalkyl,    —(CH₂)₀₋₂—R_(aryl), or —(CH₂)₀₋₂—R_(heteroaryl);-   R_(aryl) is aryl optionally substituted with one, two or three    groups independently selected from halogen, amino, mono- or    dialkylamino, —OH, —C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆ alkyl, —SO₂—N(C₁-C₆    alkyl)₂, —SO₂—(C₁-C₄ alkyl), —CO—NH₂, —CO—NH—C₁-C₆ alkyl,    —CO—N(C₁-C₆ alkyl)₂,    -   C₁-C₆ alkyl optionally substituted with one, two or three groups        independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,        —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino,    -   C₂-C₆ alkenyl or C₂-C₆ alkynyl, each of which is optionally        substituted with one, two or three groups independently selected        from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,        amino, and mono- or dialkylamino, and    -   C₁-C₆ alkoxy optionally substituted with one, two or three        groups independently selected from halogen;-   R_(heteroaryl) is heteroaryl optionally substituted with one, two or    three groups independently selected from halogen, amino, mono- or    dialkylamino, —OH, —C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆ alkyl, —SO₂—N(C₁-C₆    alkyl)₂, —SO₂—(C₁-C₄ alkyl), —CO—NH₂, —CO—NH—C₁-C₆ alkyl,    —CO—N(C₁-C₆ alkyl)₂,    -   C₁-C₆ alkyl optionally substituted with one, two or three groups        independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,        —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino,    -   C₂-C₆ alkenyl or C₂-C₆ alkynyl, each of which is optionally        substituted with one, two or three groups independently selected        from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,        amino, and mono- or dialkylamino, and    -   C₁-C₆ alkoxy optionally substituted with one, two or three        groups independently selected from halogen;-   R_(heterocyclyl) is heterocyclyl optionally substituted with one,    two or three groups independently selected from halogen, amino,    mono- or dialkylamino, —OH, —C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆ alkyl,    —SO₂—N(C₁-C₆ alkyl)₂, —SO₂—(C₁-C₄ alkyl), —CO—NH₂, —CO—NH—C₁-C₆    alkyl, ═O, —CO—N(C₁-C₆ alkyl)₂,    -   C₁-C₆ alkyl optionally substituted with one, two or three groups        independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,        —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino,    -   C₂-C₆ alkenyl or C₂-C₆ alkynyl, each of which is optionally        substituted with one, two or three groups independently selected        from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,        amino, and mono- or dialkylamino, and    -   C₁-C₆ alkoxy optionally substituted with one, two or three        groups independently selected from halogen;-   R₂ is    -   —H; or —(CH₂)₀₋₄—R_(aryl) and —(CH₂)₀₋₄—R_(heteroaryl); or    -   C₁-C₆ alkyl optionally substituted with one, two or three groups        independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,        —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino; or    -   C₂-C₆ alkenyl, C₂-C₆ alkynyl or —(CH₂)₀₋₄— C₃-C₇ cycloalkyl,        each of which is optionally substituted with one, two or three        groups independently selected from C₁-C₃ alkyl, halogen, —OH,        —SH, —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino;-   R₃ is —H, C₂-C₆ alkenyl, C₂-C₆ alkynyl, —(CH₂)₀₋₄—R_(aryl), or    —(CH₂)₀₋₄—R_(heteroaryl); or    -   C₁-C₆ alkyl optionally substituted with one, two or three groups        independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,        —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino; or    -   —(CH₂)₀₋₄—C₃-C₇ cycloalkyl optionally substituted with one, two        or three groups independently selected from C₁-C₃ alkyl,        halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or        dialkylamino; or-   R₂ and R₃ taken together with the carbon atom to which they are    attached form a carbocycle of three, four, five, six, or seven    carbon atoms, where one atom is optionally a heteroatom selected    from the group consisting of —O—, —S—, —SO₂—, and —NR₈—;-   R_(c) is hydrogen, —(CR₂₄₅R₂₅₀)₀₋₄-aryl, —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl,    —(CR₂₄₅R₂₅₀)₀₋₄-heterocyclyl, —(CR₂₄₅R₂₅₀)₀₋₄-aryl-heteroaryl,    —(CR₂₄₅R₂₅₀)₀₋₄-aryl-heterocyclyl, —(CR₂₄₅R₂₅₀)₀₋₄-aryl-aryl,    —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl-aryl,    —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl-heterocyclyl,    —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl-heteroaryl,    —(CR₂₄₅R₂₅₀)₀₋₄-heterocyclyl-heteroaryl,    —(CR₂₄₅R₂₅₀)₀₋₄-heterocyclyl-heterocyclyl,    —(CR₂₄₅R₂₅₀)₀₋₄-heterocyclyl-aryl, —[C(R₂₅₅)(R₂₆₀)]₁₋₃—CO—N—(R₂₅₅)₂,    —CH(aryl)₂, —CH(heteroaryl)₂, —CH(heterocyclyl)₂,    —CH(aryl)(heteroaryl), —(CH₂)₀₋₁—CH((CH₂)₀₋₆—OH)—(CH₂)₀₋₁-aryl,    —(CH₂)₀₋₁—CH((CH₂)₀₋₆—OH—(CH₂)₀₋₁-heteroaryl, —CH(-aryl or    -heteroaryl)-CO—O(C₁-C₄ alkyl), —CH(—CH₂—OH)—CH(OH)-phenyl-NO₂,    (C₁-C₆ alkyl)-O—(C₁-C₆ alkyl)-OH; —CH₂—NH—CH₂—CH(—O—CH₂—CH₃)₂,    —(CH₂)₀₋₆—C(═NR₂₃₅)(NR₂₃₅R₂₄₀), or    -   C₁-C₁₀ alkyl optionally substituted with 1, 2, or 3 groups        independently selected from the group consisting of R₂₀₅,        —OC═ONR₂₃₅R₂₄₀, —S(═O)₀₋₂(C₁-C₆ alkyl), —SH, —NR₂₃₅C═ONR₂₃₅R₂₄₀,        —C═ONR₂₃₅R₂₄₀, and —S(═O)₂NR₂₃₅R₂₄₀, or    -   —(CH₂)₀₋₃—(C₃-C₈) cycloalkyl wherein the cycloalkyl is        optionally substituted with 1, 2, or 3 groups independently        selected from the group consisting of R₂₀₅, —CO₂H, and        —CO₂—(C₁-C₄ alkyl), or    -   cyclopentyl, cyclohexyl, or cycloheptyl ring fused to aryl,        heteroaryl, or heterocyclyl wherein one, two or three carbons of        the cyclopentyl, cyclohexyl, or cycloheptyl is optionally        replaced with a heteroatom independently selected from NH,        NR₂₁₅, O, or S(═O)₀₋₂, and wherein the cyclopentyl, cyclohexyl,        or cycloheptyl group can be optionally substituted with one or        two groups that are independently R₂₀₅, ═O, —CO—NR₂₃₅R₂₄₀, or        —SO₂—(C₁-C₄ alkyl), or    -   C₂-C₁₀ alkenyl or C₂-C₁₀ alkynyl, each of which is optionally        substituted with 1, 2, or 3 R₂₀₅ groups, wherein    -   each aryl and heteroaryl is optionally substituted with 1, 2, or        3 R₂₀₀, and wherein each heterocyclyl is optionally substituted        with 1, 2, 3, or 4 R₂₁₀;-   R₂₀₀ at each occurrence is independently selected from —OH, —NO₂,    halogen, —CO₂H, C≡N, —(CH₂)₀₋₄—CO—NR₂₂₀R₂₂₅, —(CH₂)₀₋₄—CO—(C₁-C₁₂    alkyl), —(CH₂)₀₋₄—CO—(C₂-C₁₂ alkenyl), —(CH₂)₀₋₄—CO—(C₂-C₁₂    alkynyl), —(CH₂)₀₋₄—CO—(C₃-C₇ cycloalkyl), —(CH₂)₀₋₄—CO-aryl,    —(CH₂)₀₋₄—CO-heteroaryl, —(CH₂)₀₋₄—CO-heterocyclyl,    —(CH₂)₀₋₄—CO—O—R₂₁₅, —(CH₂)₀₋₄—SO₂—NR₂₂₀R₂₂₅, —(CH₂)₀₋₄—SO—(C₁-C₈    alkyl), —(CH₂)₀₋₄—SO₂—(C₁-C₁₂ alkyl), —(CH₂)₀₋₄—SO₂—(C₃-C₇    cycloalkyl), —(CH₂)₀₋₄—N(H or R₂₁₅)—CO—O—R₂₁₅, —(CH₂)₀₋₄—N(H or    R₂₁₅)—CO—N(R₂₁₅)₂, —(CH₂)₀₋₄—N—CS—N(R₂₁₅)₂, —(CH₂)₀₋₄—N(—H or    R₂₁₅)—CO—R₂₂₀, —(CH₂)₀₋₄—NR₂₂₀R₂₂₅, —(CH₂)₀₋₄—CO—(C₁-C₆ alkyl),    —(CH₂)₀₋₄—O—P(O)—(OR₂₄₀)₂, —(CH₂)₀₋₄—O—CO—N(R₂₁₅)₂,    —(CH₂)₀₋₄—O—CS—N(R₂₁₅)₂, —(CH₂)₀₋₄—O—(R₂₁₅),    —(CH₂)₀₋₄—O—(R₂₁₅)—COOH, —(CH₂)₀₋₄—S—(R₂₁₅), —(CH₂)₀₋₄—O—(C₁-C₆    alkyl optionally substituted with 1, 2, 3, or 5 —F), C₃-C₇    cycloalkyl, —(CH₂)₀₋₄—N(H or R₂₁₅)—SO₂—R₂₂₀, —(CH₂)₀₋₄—C₃-C₇    cycloalkyl, or    -   C₁-C₁₀ alkyl optionally substituted with 1, 2, or 3 R₂₀₅ groups,        or    -   C₂-C₁₀ alkenyl or C₂-C₁₀ alkynyl, each of which is optionally        substituted with 1 or 2 R₂₀₅ groups, wherein    -   the aryl and heteroaryl groups at each occurrence are optionally        substituted with 1, 2, or 3 groups that are independently R₂₀₅,        R₂₁₀, or        -   C₁-C₆ alkyl substituted with 1, 2, or 3 groups that are            independently R₂₀₅ or R₂₁₀, and wherein    -   the heterocyclyl group at each occurrence is optionally        substituted with 1, 2, or 3 groups that are independently R₂₁₀;-   R₂₀₅ at each occurrence is independently selected from C₁-C₆ alkyl,    halogen, —OH, —O-phenyl, —SH, —C≡N, —CF₃, C₁-C₆ alkoxy, NH₂,    NH(C₁-C₆ alkyl) or N—(C₁-C₆ alkyl) (C₁-C₆ alkyl);-   R₂₁₀ at each occurrence is independently selected from halogen,    C₁-C₆ alkoxy, C₁-C₆ haloalkoxy, —NR₂₂₀R₂₂₅, OH, C≡N, —CO—(C₁-C₄    alkyl), —SO₂—NR₂₃₅R₂₄₀, —CO—NR₂₃₅R₂₄₀, —SO₂—(C₁-C₄ alkyl), ═O, or    -   C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl or C₃-C₇ cycloalkyl,        each of which is optionally substituted with 1, 2, or 3 R₂₀₅        groups;-   R₂₁₅ at each occurrence is independently selected from C₁-C₆ alkyl,    —(CH₂)₀₋₂-(aryl), C₂-C₆ alkenyl, C₂-C₆ alkynyl, C₃-C₇ cycloalkyl,    and —(CH₂)₀₋₂-(heteroaryl), —(CH₂)₀₋₂-(heterocyclyl), wherein    -   the aryl group at each occurrence is optionally substituted with        1, 2, or 3 groups that are independently R₂₀₅ or R₂₁₀, and        wherein    -   the heterocyclyl and heteroaryl groups at each occurrence are        optionally substituted with 1, 2, or 3 R₂₁₀;-   R₂₂₀ and R₂₂₅ at each occurrence are independently selected from —H,    —C₃-C₇ cycloalkyl, —(C₁-C₂ alkyl)-(C₃-C₇ cycloalkyl), —(C₁-C₆    alkyl)-O—(C₁-C₃ alkyl), —C₂-C₆ alkenyl, —C₂-C₆ alkynyl, —C₁-C₆ alkyl    chain with one double bond and one triple bond, -aryl, -heteroaryl,    and -heterocyclyl, or    -   —C₁-C₁₀ alkyl optionally substituted with —OH, —NH₂ or halogen,        wherein    -   the aryl, heterocyclyl and heteroaryl groups at each occurrence        are optionally substituted with 1, 2, or 3 R₂₇₀ groups-   R₂₃₅ and R₂₄₀ at each occurrence are independently H, or C₁-C₆    alkyl;-   R₂₄₅ and R₂₅₀ at each occurrence are independently selected from —H,    C₁-C₄ alkyl, C₁-C₄ alkylaryl, C₁-C₄ alkylheteroaryl, C₁-C₄    hydroxyalkyl, C₁-C₄ alkoxy, C₁-C₄ haloalkoxy, —(CH₂)₀₋₄—C₃-C₇    cycloalkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, and phenyl; or-   R₂₄₅ and R₂₅₀ are taken together with the carbon to which they are    attached to form a carbocycle of 3, 4, 5, 6, or 7 carbon atoms,    where one carbon atom is optionally replaced by a heteroatom    selected from —O—, —S—, —SO₂—, and —NR₂₂₀—;-   R₂₅₅ and R₂₆₀ at each occurrence are independently selected from —H,    —(CH₂)₁₋₂—S(O)₀₋₂—(C₁-C₆ alkyl), —(C₁-C₄ alkyl)-aryl, —(C₁-C₄    alkyl)-heteroaryl, —(C₁-C₄ alkyl)-heterocyclyl, -aryl, -heteroaryl,    -heterocyclyl, —(CH₂)₁₋₄—R₂₆₅—(CH₂)₀₋₄-aryl,    —(CH₂)₁₋₄—R₂₆₅—(CH₂)₀₋₄-heteroaryl,    —(CH₂)₁₋₄—R₂₆₅—(CH₂)₀₋₄-heterocyclyl, or    -   C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl or —(CH₂)₀₋₄—C₃-C₇        cycloalkyl, each of which is optionally substituted with 1, 2,        or 3 R₂₀₅ groups, wherein    -   each aryl or phenyl is optionally substituted with 1, 2, or 3        groups that are independently R₂₀₅, R₂₁₀, or        -   C₁-C₆ alkyl substituted with 1, 2, or 3 groups that are            independently R₂₀₅ or R₂₁₀, and wherein    -   each heterocyclyl is optionally substituted with 1, 2, 3, or 4        R₂₁₀;-   R₂₆₅ at each occurrence is independently —O—, —S— or —N(C₁-C₆    alkyl)-;-   R₂₇₀ at each occurrence is independently R₂₀₅, halogen C₁-C₆ alkoxy,    C₁-C₆ haloalkoxy, NR₂₃₅R₂₄₀, —OH, —C≡N, —CO—(C₁-C₄ alkyl),    —SO₂—NR₂₃₅R₂₄₀, —CO—NR₂₃₅R₂₄₀, —SO₂—(C₁-C₄ alkyl), ═O, or    -   C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl or —(CH₂)₀₋₄—C₃-C₇        cycloalkyl, each of which is optionally substituted with 1, 2,        or 3 R₂₀₅ groups;-   and pharmaceutically acceptable salts thereof.

The invention also provides intermediates and methods useful forpreparing the compounds of formula X.

The invention further provides pharmaceutical compositions comprising acompound of formula X.

The present invention also provides the use of a compound of formula (X)and pharmaceutically acceptable salts thereof for the manufacture of amedicament.

The present invention also provides a method of treating a patient whohas Alzheimer's Disease or other diseases that can be treated byinhibiting beta-secretase activity.

DETAILED DESCRIPTION OF THE INVENTION

The compounds encompassed by the instant invention are those describedby the general formula (IX)set forth above, and the pharmaceuticallyacceptable salts and prodrugs thereof.

In an embodiment, the compounds of formula (IX) have synstereochemistry.

In an embodiment, compounds of formula (IX) have anti stereochemistry.

The invention also provides intermediates and methods useful forpreparing the compounds of formula IX.

In an embodiment, the compounds of the invention have the formula (IXa):

where R, X, Y, R₂, R₃, R₆ and R_(c) are as defined above for (IX).Preferred compounds of formula (IXa) are those in which Y is alkynyl orY together with the carbon to which it is attached is a D or L aminoacid side chain; X is C₁-C₆ alkyl; R₂ and R₃ are hydrogen; and R_(c) is—(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXb):

where R, X, B, R₂, R₃ and R_(c) are as defined above for (IX). Preferredcompounds of formula (IXb) are those in which X is C₁-C₆ alkyl; B isaryl optionally substituted with R₆; R₂ and R₃ are hydrogen; and R_(c)is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXc):

where R, X, B, R₂, R₃ and R_(c) are as defined above for (IX) Preferredcompounds of formula (IXc) are those in which X is C₁-C₆ alkyl; B isaryl optionally substituted with R₆; R₂ and R₃ are hydrogen; and R_(c)is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXd):

where R, X, B, R₂, R₃ and R_(c) are as defined above for (IX). Preferredcompounds of formula (IXd) are those in which X is C₁-C₆ alkyl; B isaryl optionally substituted with R₆; R₂ and R₃ are hydrogen; and R_(c)is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXe):

where R, X, B, R₂, R₃ and R_(c) are as defined above for (IX). Preferredcompounds of formula (IXe) are those in which X is C₁-C₆ alkyl; B isaryl optionally substituted with R₆; R₂ and R₃ are hydrogen; and R_(c)is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXf):

where Y, X, B, R₂, R₃ and R_(c) are as defined above for (IX). Preferredcompounds of formula (IXf) are those in which X is C₁-C₆ alkyl; Y isalkynyl or Y together with the carbon to which it is attached is a D orL amino acid side chain; B is aryl optionally substituted with R₆; R₂and R₃ are hydrogen; and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which is optionally substituted withone or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXg):

where Y, X, B, R₂, R₃ and R_(c) are as defined above for (IX). Preferredcompounds of formula (IXg) are those in which X is C₁-C₆ alkyl; Y isalkynyl or Y together with the carbon to which it is attached is a D orL amino acid side chain; B is aryl optionally substituted with R₆; R₂and R₃ are hydrogen; and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which is optionally substituted withone or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXh):

where Y, B, R₂, R₃ and R_(c) are as defined above for (IX). Preferredcompounds of formula (IXh) are those in which Y is alkynyl or Y togetherwith the carbon to which it is attached is a D or L amino acid sidechain; B is aryl optionally substituted with R₆; R₂ and R₃ are hydrogen;and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each ofwhich is optionally substituted with one or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXi):

where Y, X, B, R and R_(c) are as defined above for (IX). Preferredcompounds of formula (IXi) are those in which X is C₁-C₆ alkyl; Y isalkynyl or Y together with the carbon to which it is attached is a D orL amino acid side chain; B is aryl optionally substituted with R₆; andR_(c) is —(CR₂₄₅R₂₅₀)₀ ₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each ofwhich is optionally substituted with one or two R₂₀₀.

In another embodiment, the compounds of the invention have the formula(IXj):

where Y, X, B, R and aryl are as defined above for (IX). Preferredcompounds of formula (IXj) are those in which X is C₁-C₆ alkyl; Y isalkynyl or Y together with the carbon to which it is attached is a D orL amino acid side chain; B is aryl optionally substituted with R₆; andR₂₀₀ is C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkenyl, trifluoromethyl, orhalogen.

In another embodiment, the compounds of the invention have the formula(IXk):

where Y, X, B, R₂, R₃ and R₂₀₀ are as defined above for (IX), and V isCH or N. Preferred compounds of formula (IXk) are those in which X isC₁-C₆ alkyl; Y is alkynyl or Y together with the carbon to which it isattached is a D or L amino acid side chain; B is aryl optionallysubstituted with R₆; R₂ and R₃ are hydrogen; and R₂₀₀ is C₁-C₆ alkyl,C₂-C₆ alkenyl, C₂-C₆ alkenyl, trifluoromethyl, or halogen.

In another embodiment, the compounds of the invention have the formula(IXl):

where Y, X, B, R, R₂ and R₃ are as defined above for (IX). Preferredcompounds of formula (IXl) are those in which X is C₁-C₆ alkyl; Y isalkynyl or Y together with the carbon to which it is attached is a D orL amino acid side chain; B is aryl optionally substituted with R₆; andR₂ and R₃ are hydrogen.

In another embodiment, the compounds of the invention have the formula(IXm):

where Y, X, B, R, R₂ and R₃ are as defined above for (IX). Preferredcompounds of formula (IXm) are those in which X is C₁-C₆ alkyl; Y isalkynyl or Y together with the carbon to which it is attached is a D orL amino acid side chain; B is aryl optionally substituted with R₆; andR₂ and R₃ are hydrogen.

In another embodiment, the compounds of the invention have the formula(IXn):

where Y, X, B, R, R₂ and R₃ are as defined above for (IX). Preferredcompounds of formula (IXn) are those in which X is C₁-C₆ alkyl; Y isalkynyl or Y together with the carbon to which it is attached is a D orL amino acid side chain; B is aryl optionally substituted with R₆; andR₂ and R₃ are hydrogen.

In an embodiment, the compound of formula (IX) includes apharmaceutically acceptable salt selected from the group consisting ofsalts of the following acids: hydrochloric, hydrobromic, hydroiodic,nitric, sulfuric, phosphoric, citric, TFA, methanesulfonic,CH₃—(CH₂)_(n)—COOH where n is 0 thru 4, HOOC—(CH₂) n-COOH where n is asdefined above, HOOC—CH═CH—COOH, and phenyl-COOH.

The present invention also includes compounds of the formula (II):

where X and B are as defined above and R_(x) is a suitable organiccarboxylic acid derivative-functional group; or a chemically acceptablesalt thereof. In a preferred embodiment, R_(x) is C₁-C₆ alkyl.

The present invention also includes an alcohol of formula (III):

where X, B, R₂, R₃, and R_(x) are as defined above and X1 X₁ is aleaving group including, but not limited to, —Cl, —Br, —I, —O-tosylate,—O-mesylate, —O-nosylate; or a chemically acceptable salt thereof. In apreferred embodiment, R_(x) is C₁-C₆ alkyl.

In an embodiment, this alcohol includes as Protecting Groupt-butoxycarbonyl.

In an embodiment, this alcohol includes as Protecting Groupbenzyloxycarbonyl.

In an embodiment, this alcohol includes as X₁ —Cl or —Br.

The present invention also includes an epoxide of the formula (IV):

where X, B, R₂, R₃, and R_(x) are as defined above; or a chemicallyacceptable salt thereof. In a preferred embodiment, R_(x) is C₁-C₆alkyl.

In an embodiment, this epoxide includes as Protecting Groupt-butoxycarbonyl.

In an embodiment, this epoxide includes as Protecting Groupbenzyloxycarbonyl.

The present invention also includes a compound of formula (VI):

where X, B, R₂, R₃, R_(c), and R_(x) are as defined above; or achemically acceptable salt thereof. In a preferred embodiment, R_(x) isC₁-C₆ alkyl.

In an embodiment, this protected alcohol includes as Protecting Groupt-butoxycarbonyl.

In an embodiment, this protected alcohol includes as Protecting Groupbenzyloxycarbonyl.

The present invention also includes an amine of formula (VII):

where X, B, R₂, R₃, R_(x) and R_(c) are as defined above; or achemically acceptable salt thereof. In a preferred embodiment, R_(x) isC₁-C₆ alkyl.

The present invention also includes an amine of formula (VIII):

The present invention also includes a method of treating a patient whohas, or in preventing a patient from getting, a disease or conditionselected from the group consisting of Alzheimer's disease, for helpingprevent or delay the onset of Alzheimer's disease, for treating patientswith mild cognitive impairment (MCI) and preventing or delaying theonset of Alzheimer's disease in those who would progress from MCI to AD,for treating Down's syndrome, for treating humans who have HereditaryCerebral Hemorrhage with Amyloidosis of the Dutch-Type, for treatingcerebral amyloid angiopathy and preventing its potential consequences,i.e. single and recurrent lobar hemorrhages, for treating otherdegenerative dementias, including dementias of mixed vascular anddegenerative origin, dementia associated with Parkinson's disease,dementia associated with progressive supranuclear palsy, dementiaassociated with cortical basal degeneration, or diffuse Lewy body typeof Alzheimer's disease and who is in need of such treatment whichincludes administration of a therapeutically effective amount of acompound of formula (X) and pharmaceutically acceptable salts thereof.

In an embodiment, this method of treatment can be used where the diseaseis Alzheimer's disease.

In an embodiment, this method of treatment can help prevent or delay theonset of Alzheimer's disease.

In an embodiment, this method of treatment can be used where the diseaseis mild cognitive impairment.

In an embodiment, this method of treatment can be used where the diseaseis Down's syndrome.

In an embodiment, this method of treatment can be used where the diseaseis Hereditary Cerebral Hemorrhage with Amyloidosis of the Dutch-Type.

In an embodiment, this method of treatment can be used where the diseaseis cerebral amyloid angiopathy.

In an embodiment, this method of treatment can be used where the diseaseis degenerative dementias.

In an embodiment, this method of treatment can be used where the diseaseis diffuse Lewy body type of Alzheimer's disease.

In an embodiment, this method of treatment can treat an existingdisease.

In an embodiment, this method of treatment can prevent a disease fromdeveloping.

In an embodiment, this method of treatment can employ therapeuticallyeffective amounts: for oral administration from about 0.1 mg/day toabout 1,000 mg/day; for parenteral, sublingual, intranasal, intrathecaladministration from about 0.5 to about 100 mg/day; for depoadministration and implants from about 0.5 mg/day to about 50 mg/day;for topical administration from about 0.5 mg/day to about 200 mg/day;for rectal administration from about 0.5 mg to about 500 mg.

In an embodiment, this method of treatment can employ therapeuticallyeffective amounts: for oral administration from about 1 mg/day to about100 mg/day; and for parenteral administration from about 5 to about 50mg daily.

In an embodiment, this method of treatment can employ therapeuticallyeffective amounts for oral administration from about 5 mg/day to about50 mg/day.

The present invention also includes a pharmaceutical composition whichincludes a compound of formula (IX) and pharmaceutically acceptablesalts thereof.

The present invention also includes the use of a compound of formula(IX) and pharmaceutically acceptable salts thereof for the manufactureof a medicament for use in treating a patient who has, or in preventinga patient from getting, a disease or condition selected from the groupconsisting of Alzheimer's disease, for helping prevent or delay theonset of Alzheimer's disease, for treating patients with mild cognitiveimpairment (MCI) and preventing or delaying the onset of Alzheimer'sdisease in those who would progress from MCI to AD, for treating Down'ssyndrome, for treating humans who have Hereditary Cerebral Hemorrhagewith Amyloidosis of the Dutch-Type, for treating cerebral amyloidangiopathy and preventing its potential consequences, i.e. single andrecurrent lobar hemorrhages, for treating other degenerative dementias,including dementias of mixed vascular and degenerative origin, dementiaassociated with Parkinson's disease, dementia associated withprogressive supranuclear palsy, dementia associated with cortical basaldegeneration, diffuse Lewy body type of Alzheimer's disease and who isin need of such treatment.

In an embodiment, this use of a compound of formula (IX) can be employedwhere the disease is Alzheimer's disease.

In an embodiment, this use of a compound of formula (IX) can helpprevent or delay the onset of Alzheimer's disease.

In an embodiment, this use of a compound of formula (IX) can be employedwhere the disease is mild cognitive impairment.

In an embodiment, this use of a compound of formula (IX) can be employedwhere the disease is Down's syndrome.

In an embodiment, this use of a compound of formula (IX) can be employedwhere the disease is Hereditary Cerebral Hemorrhage with Amyloidosis ofthe Dutch-Type.

In an embodiment, this use of a compound of formula (IX) can be employedwhere the disease is cerebral amyloid angiopathy.

In an embodiment, this use of a compound of formula (IX) can be employedwhere the disease is degenerative dementias.

In an embodiment, this use of a compound of formula (IX) can be employedwhere the disease is diffuse Lewy body type of Alzheimer's disease.

In an embodiment, this use of a compound employs a pharmaceuticallyacceptable salt selected from the group consisting of salts of thefollowing acids hydrochloric, hydrobromic, hydroiodic, nitric, sulfuric,phosphoric, citric, TFA, methanesulfonic, CH₃—(CH₂)_(n)—COOH where n is0 thru 4, HOOC—(CH₂) n-COOH where n is as defined above,HOOC—CH═CH—COOH, and phenyl-COOH.

The present invention also includes methods for inhibitingbeta-secretase activity, for inhibiting cleavage of amyloid precursorprotein (APP), in a reaction mixture, at a site between Met596 andAsp597, numbered for the APP-695 amino acid isotype, or at acorresponding site of an isotype or mutant thereof; for inhibitingproduction of amyloid beta peptide (A beta) in a cell; for inhibitingthe production of beta-amyloid plaque in an animal; and for treating orpreventing a disease characterized by beta-amyloid deposits in the brainwhich include administration of a therapeutically effective amount of acompound of formula (IX) and pharmaceutically acceptable salts thereof.

The present invention also includes a method for inhibitingbeta-secretase activity, including exposing said beta-secretase to aneffective inhibitory amount of a compound of the formula (IX) or apharmaceutically acceptable salt thereof.

Preferably, this method employs a compound that inhibits 50% of theenzyme's activity at a concentration of less than 50 micromolar.

This method more preferably employs a compound that inhibits 50% of theenzyme's activity at a concentration of 10 micromolar or less.

This method even more preferably employs a compound that inhibits 50% ofthe enzyme's activity at a concentration of 1 micromolar or less.

In a particular embodiment, this method employs a compound that inhibits50% of the enzyme's activity at a concentration of 10 nanomolar or less.

In an embodiment, this method includes exposing said beta-secretase tosaid compound in vitro.

In an embodiment, this method includes exposing said beta-secretase tosaid compound in a cell.

In an embodiment, this method includes exposing said beta-secretase tosaid compound in a cell in an animal.

In an embodiment, this method includes exposing said beta-secretase tosaid compound in a human.

The present invention also includes a method for inhibiting cleavage ofamyloid precursor protein (APP), in a reaction mixture, at a sitebetween Met596 and Asp597, numbered for the APP-695 amino acid isotype;or at a corresponding site of an isotype or mutant thereof, includingexposing said reaction mixture to an effective inhibitory amount of acompound of formula (IX) or a pharmaceutically acceptable salt thereof.

In an embodiment, this method employs a cleavage site: between Met652and Asp653, numbered for the APP-751 isotype; between Met 671 and Asp672, numbered for the APP-770 isotype; between Leu596 and Asp597 of theAPP-695 Swedish Mutation; between Leu652 and Asp653 of the APP-751Swedish Mutation; or between Leu671 and Asp672 of the APP-770 SwedishMutation.

In an embodiment, this method exposes said reaction mixture in vitro.

In an embodiment, this method exposes said reaction mixture in a cell.

In an embodiment, this method exposes said reaction mixture in an animalcell.

In an embodiment, this method exposes said reaction mixture in a humancell.

The present invention also includes a method for inhibiting productionof amyloid beta peptide (A beta) in a cell, including administering tosaid cell an effective inhibitory amount of a compound of formula (IX)or a pharmaceutically acceptable salt thereof.

In an embodiment, this method includes administering to an animal.

In an embodiment, this method includes administering to a human.

The present invention also includes a method for inhibiting theproduction of beta-amyloid plaque in an animal, including administeringto said animal an effective inhibitory amount of a compound of theformula (IX) or a pharmaceutically acceptable salt thereof.

In an embodiment, this method includes administering to a human.

The present invention also includes a method for treating or preventinga disease characterized by beta-amyloid deposits in the brain includingadministering to a patient an effective therapeutic amount of ahydroxyethylene compound of the formula (IX) or a pharmaceuticallyacceptable salt thereof.

In an embodiment, this method employs a compound that inhibits 50% ofthe enzyme's activity at a concentration of less than 50 micromolar.

In an embodiment, this method employs a compound that inhibits 50% ofthe enzyme's activity at a concentration of 10 micromolar or less.

In an embodiment, this method employs a compound that inhibits 50% ofthe enzyme's activity at a concentration of 1 micromolar or less.

In an embodiment, this method employs a compound that inhibits 50% ofthe enzyme's activity at a concentration of 10 nanomolar or less.

In an embodiment, this method employs a compound at a therapeutic amountin the range of from about 0.1 to about 1000 mg/day.

In an embodiment, this method employs a compound at a therapeutic amountin the range of from about 15 to about 1500 mg/day.

In an embodiment, this method employs a compound at a therapeutic amountin the range of from about 1 to about 100 mg/day.

In an embodiment, this method employs a compound at a therapeutic amountin the range of from about 5 to about 50 mg/day.

In an embodiment, this method can be used where said disease isAlzheimer's disease.

In an embodiment, this method can be used where said disease is MildCognitive Impairment, Down's Syndrome, or Hereditary Cerebral Hemorrhagewith Amyloidosis of the Dutch Type.

The present invention also includes a composition includingbeta-secretase complexed with a compound of formula (IX) or apharmaceutically acceptable salt thereof.

The present invention also includes a method for producing abeta-secretase complex including exposing beta-secretase to a compoundof formula (IX) or a pharmaceutically acceptable salt thereof, in areaction mixture under conditions suitable for the production of saidcomplex.

In an embodiment, this method employs exposing in vitro.

In an embodiment, this method employs a reaction, mixture that is acell.

The present invention also includes a component kit including componentparts capable of being assembled, in which at least one component partincludes a compound of formula Xa enclosed in a container.

In an embodiment, this component kit includes lyophilized compound, andat least one further component part includes a diluent.

The present invention also includes a container kit including aplurality of containers, each container including one or more unit doseof a compound of formula (IX) or a pharmaceutically acceptable saltthereof.

In an embodiment, this container kit includes each container adapted fororal delivery and includes a tablet, gel, or capsule.

In an embodiment, this container kit includes each container adapted forparenteral delivery and includes a depot product, syringe, ampoule, orvial.

In an embodiment, this container kit includes each container adapted fortopical delivery and includes a patch, medipad, ointment, or cream.

The present invention also includes an agent kit including a compound offormula (IX) or a pharmaceutically acceptable salt thereof; and one ormore therapeutic agent selected from the group consisting of anantioxidant, an anti-inflammatory, a gamma secretase inhibitor, aneurotrophic agent, an acetyl cholinesterase inhibitor, a statin, an Abeta peptide, and an anti-A beta antibody.

The present invention also includes a composition including a compoundof formula (IX) or a pharmaceutically acceptable salt thereof; and aninert diluent or edible carrier.

In an embodiment, this composition includes a carrier that is an oil.

The present invention also includes a composition including a compoundof formula (IX) or a pharmaceutically acceptable salt thereof; and abinder, excipient, disintegrating agent, lubricant, or gildant.

The present invention also includes a composition including a compoundof formula (IX) or a pharmaceutically acceptable salt thereof; disposedin a cream, ointment, or patch.

The present invention provides compounds, compositions, kits, andmethods for inhibiting beta-secretase-mediated cleavage of amyloidprecursor protein (APP). More particularly, the compounds, compositions,and methods of the invention are effective to inhibit the production ofA beta peptide and to treat or prevent any human or veterinary diseaseor condition associated with a pathological form of A beta peptide.

The compounds, compositions, and methods of the invention are useful fortreating humans who have Alzheimer's Disease (AD), for helping preventor delay the onset of AD, for treating patients with mild cognitiveimpairment (MCI), and preventing or delaying the onset of AD in thosepatients who would otherwise be expected to progress from MCI to AD, fortreating Down's syndrome, for treating Hereditary Cerebral Hemorrhagewith Amyloidosis of the Dutch Type, for treating cerebral beta-amyloidangiopathy and preventing its potential consequences such as single andrecurrent lobar hemorrhages, for treating other degenerative dementias,including dementias of mixed vascular and degenerative origin, fortreating dementia associated with Parkinson's disease, dementiaassociated with progressive supranuclear palsy, dementia associated withcortical basal degeneration, and diffuse Lewy body type AD.

The compounds of the invention possess beta-secretase inhibitoryactivity. The inhibitory activities of the compounds of the inventionare readily demonstrated, for example, using one or more of the assaysdescribed herein or known in the art.

By “Protecting Group” in the present invention is meant any suitableorganic protecting group such as disclosed in T. W. Green and P. G. M.Wuts in “Protective Groups in Organic Chemistry, John Wiley and Sons,1991. Preferred protecting groups in the present invention aret-butoxycarbonyl, benzyloxycarbonyl, formyl, trityl, phthalimido,trichloroacetyl, chloroacetyl, bromoacetyl, iodoacetyl,4-phenylbenzyloxycarbonyl, 2-methylbenzyloxycarbonyl,4-ethoxybenzyloxycarbonyl, 4-fluorobenzyloxycarbonyl,4-chlorobenzyloxycarbonyl, 3-chlorobenzyloxycarbonyl,2-chlorobenzyloxycarbonyl, 2,4-dichlorobenzyloxycarbonyl,4-bromobenzyloxycarbonyl, 3-bromobenzyloxycarbonyl,4-nitrobenzyloxycarbonyl, 4-cyanobenzyloxycarbonyl, 2-(4-xenyl)isopropoxycarbonyl, 1,1-diphenyleth-1-yloxycarbonyl,1,1-diphenylprop-1-yloxycarbonyl, 2-phenylprop-2-yloxycarbonyl,2-(p-toluyl)prop-2-yloxycarbonyl, cyclopentanyloxycarbonyl,1-methylcycoopentanyloxycarbonyl, cyclohexanyloxycarbonyl,1-methylcyclohexanyloxycabonyl, 2-methylcyclohexanyloxycarbonyl,2-(4-toluylsulfonyl)ethoxycarbonyl, 2-(methylsulfonyl)ethoxycarbonyl,2-(triphenylphosphino)ethoxycarbonyl, fluorenylmethoxycarbonyl,2-(trimethylsilyl) ethoxycarbonyl, allyloxycarbonyl,1-(trimethylsilylmethyl) prop-1-enyloxycarbonyl,5-benzisoxalylmethoxycarbonyl, 4-acetoxybenzyloxycarbonyl,2,2,2-trichloroethoxycarbonyl, 2-ethynyl-2-propoxycarbonyl,cyclopropylmethoxycarbonyl, 4-(decyloxyl)benzyloxycarbonyl,isobrornyloxycarbonyl, 1-piperidyloxycarbonyl, 9-fluoroenylmethylcarbonate, —CH—CH═CH₂, or phenyl-C(═N—)—H.

By “alkyl” and “C₁-C₆ alkyl” in the present invention is meant straightor branched chain alkyl groups having 1-6 carbon atoms, such as, methyl,ethyl, propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, pentyl,2-pentyl, isopentyl, neopentyl, hexyl, 2-hexyl, 3-hexyl, and3-methylpentyl. It is understood that in cases where an alkyl chain of asubstituent (e.g. of an alkyl, alkoxy or alkenyl group) is shorter orlonger than 6 carbons, it will be so indicated in the second “C” as, forexample, “C₁-C₁₀” indicates a maximum of 10 carbons.

By “alkoxy” and “C₁-C₆ alkoxy” in the present invention is meantstraight or branched chain alkyl groups having 1-6 carbon atoms,attached through at least one divalent oxygen atom, such as, forexample, methoxy, ethoxy, propoxy, isopropoxy, n-butoxy, sec-butoxy,tert-butoxy, pentoxy, isopentoxy, neopentoxy, hexoxy, and3-methylpentoxy.

By the term “halogen” in the present invention is meant fluorine,bromine, chlorine, and iodine.

“Alkenyl” and “C₂-C₆ alkenyl” means straight and branched hydrocarbonradicals having from 2 to 6 carbon atoms and from one to three doublebonds and includes, for example, ethenyl, propenyl, 1-but-3-enyl,1-pent-3-enyl, 1-hex-5-enyl and the like.

“Alkynyl” and “C₂-C₆ alkynyl” means straight and branched hydrocarbonradicals having from 2 to 6 carbon atoms and one or two triple bonds andincludes ethynyl, propynyl, butynyl, pentyn-2-yl and the like.

As used herein, the term “cycloalkyl” refers to saturated carbocyclicradicals having three to twelve carbon atoms. The cycloalkyl can bemonocyclic, or a polycyclic fused system. Examples of such radicalsinclude cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl andcycloheptyl. The cycloalkyl groups herein are unsubstituted or, asspecified, substituted in one or more substitutable positions withvarious groups. For example, such cycloalkyl groups may be optionallysubstituted with C₁-C₆ alkyl, C₁-C₆ alkoxy, halogen, hydroxy, cyano,nitro, amino, mono(C₁-C₆)alkylamino, di(C₁-C₆)alkylamino, C₂-C₆alkenyl,C₂-C₆alkynyl, C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, amino(C₁-C₆)alkyl,mono(C₁-C₆)alkylamino (C₁-C₆)alkyl or di(C₁-C₆)alkylamino(C₁-C₆)alkyl.

By “aryl” is meant an aromatic carbocyclic group having a single ring(e.g., phenyl), multiple rings (e.g., biphenyl), or multiple condensedrings in which at least one is aromatic, (e.g.,1,2,3,4-tetrahydronaphthyl, naphthyl), which is optionally mono-, di-,or trisubstituted. Preferred aryl groups of the present invention arephenyl, 1-naphthyl, 2-naphthyl, indanyl, indenyl, dihydronaphthyl,tetralinyl or 6,7,8,9-tetrahydro-5H-benzo[a]cycloheptenyl. The arylgroups herein are unsubstituted or, as specified, substituted in one ormore substitutable positions with various groups. For example, such arylgroups may be optionally substituted with, for example, C₁-C₆ alkyl,C₁-C₆ alkoxy, halogen, hydroxy, cyano, nitro, amino,mono(C₁-C₆)alkylamino, di(C₁-C₆)alkylamino, C₂-C₆alkenyl, C₂-C₆alkynyl,C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, amino(C₁-C₆)alkyl, mono(C₁-C₆)alkylamino(C₁-C₆)alkyl, di(C₁-C₆)alkylamino(C₁-C₆)alkyl, —COOH,—C(═O)O(C₁-C₆ alkyl), —C(═O)NH₂, —C(═O)N(mono- or di-C₁-C₆ alkyl),—S(C₁-C₆ alkyl), —SO₂(C₁-C₆ alkyl), —O—C(═O)(C₁-C₆ alkyl),—NH—C(═O)—(C₁-C₆ alkyl), —N(C₁-C₆ alkyl)-C(═O)—(C₁-C₆alkyl),—NH—SO₂—(C₁-C₆ alkyl), —N(C₁-C₆ alkyl)-SO₂—(C₁-C₆ alkyl),—NH—C(═O) NH₂, —NH—C(═O)N(mono- or di-C₁-C₆ alkyl), —NH(C₁-C₆alkyl)-C(═O)—NH₂ or —NH(C₁-C₆ alkyl)-C(═O)—N-(mono- or di-C₁-C₆ alkyl).

By “heteroaryl” is meant one or more aromatic ring systems of 5-, 6-, or7-membered rings which includes fused ring systems of 9-11 atomscontaining at least one and up to four heteroatoms selected fromnitrogen, oxygen, or sulfur. Preferred heteroaryl groups of the presentinvention include pyridinyl, pyrimidinyl, quinolinyl, benzothienyl,indolyl, indolinyl, pryidazinyl, pyrazinyl, isoindolyl, isoquinolyl,quinazolinyl, quinoxalinyl, phthalazinyl, imidazolyl, isoxazolyl,pyrazolyl, oxazolyl, thiazolyl, indolizinyl, indazolyl, benzothiazolyl,benzimidazolyl, benzofuranyl, furanyl, thienyl, pyrrolyl, oxadiazolyl,thiadiazolyl, triazolyl, tetrazolyl, oxazolopyridinyl, imidazopyridinyl,isothiazolyl, naphthyridinyl, cinnolinyl, carbazolyl, beta-carbolinyl,isochromanyl, chromanyl, tetrahydroisoquinolinyl, isoindolinyl,isobenzotetrahydrofuranyl, isobenzotetrahydrothienyl, isobenzothienyl,benzoxazolyl, pyridopyridinyl, benzotetrahydrofuranyl,benzotetrahydrothienyl, purinyl, benzodioxolyl, triazinyl, phenoxazinyl,phenothiazinyl, pteridinyl, benzothiazolyl, imidazopyridinyl,imidazothiazolyl, dihydrobenzisoxazinyl, benzisoxazinyl, benzoxazinyl,dihydrobenzisothiazinyl, benzopyranyl, benzothiopyranyl, coumarinyl,isocoumarinyl, chromonyl, chromanonyl, pyridinyl-N-oxide,tetrahydroquinolinyl, dihydroquinolinyl, dihydroquinolinonyl,dihydroisoquinolinonyl, dihydrocoumarinyl, dihydroisocoumarinyl,isoindolinonyl, benzodioxanyl, benzoxazolinonyl, pyrrolyl N-oxide,pyrimidinyl N-oxide, pyridazinyl N-oxide, pyrazinyl N-oxide, quinolinylN-oxide, indolyl N-oxide, indolinyl N-oxide, isoquinolyl N-oxide,quinazolinyl N-oxide, quinoxalinyl N-oxide, phthalazinyl N-oxide,imidazolyl N-oxide, isoxazolyl N-oxide, oxazolyl N-oxide, thiazolylN-oxide, indolizinyl N-oxide, indazolyl N-oxide, benzothiazolyl N-oxide,benzimidazolyl N-oxide, pyrrolyl N-oxide, oxadiazolyl N-oxide,thiadiazolyl N-oxide, triazolyl N-oxide, tetrazolyl N-oxide,benzothiopyranyl S-oxide, benzothiopyranyl S,S-dioxide. The heteroarylgroups herein are unsubstituted or, as specified, substituted in one ormore substitutable positions with various groups. For example, suchheteroaryl groups may be optionally substituted with C₁-C₆ alkyl, C₁-C₆alkoxy, halogen, hydroxy, cyano, nitro, amino, mono(C₁-C₆)alkylamino,di(C₁-C₆)alkylamino, C₂-C₆alkenyl, C₂-C₆alkynyl, C₁-C₆ haloalkyl, C₁-C₆haloalkoxy, amino(C₁-C₆)alkyl, mono(C₁-C₆)alkylamino(C₁-C₆)alkyl ordi(C₁-C₆)alkylamino(C₁-C₆)alkyl, —COOH, —C(═O)O(C₁-C₆ alkyl), —C(═O)NH₂,—C(═O)N(mono- or di-C₁-C₆ alkyl), —S(C₁-C₆ alkyl), —SO₂(C₁-C₆ alkyl),—O—C(═O)(C₁-C₆ alkyl), —NH—C(═O)—(C₁-C₆ alkyl), —N(C₁-C₆alkyl)-C(═O)—(C₁-C₆ alkyl), —NH—SO₂—(C₁-C₆ alkyl), —N(C₁-C₆alkyl)-SO₂—(C₁-C₆ alkyl), —NH—C(═O)NH₂, —NH—C(═O)N(mono- or di-C₁-C₆alkyl), —NH(C₁-C₆ alkyl)-C(═O)—NH₂ or —NH(C₁-C₆ alkyl)-C(═O)—N-(mono- ordi-C₁-C₆ alkyl).

By “heterocycle”, “heterocycloalkyl” or “heterocyclyl” is meant one ormore carbocyclic ring systems of 4-, 5-, 6-, or 7-membered rings whichincludes fused ring systems of 9-11 atoms containing at least one and upto four heteroatoms selected from nitrogen, oxygen, or sulfur. Preferredheterocycles of the present invention include morpholinyl,thiomorpholinyl, thiomorpholinyl S-oxide, thiomorpholinyl S,S-dioxide,piperazinyl, homopiperazinyl, pyrrolidinyl, pyrrolinyl,tetrahydropyranyl, piperidinyl, tetrahydrofuranyl, tetrahydrothienyl,homopiperidinyl, homomorpholinyl, homothiomorpholinyl,homothiomorpholinyl S,S-dioxide, oxazolidinonyl, dihydropyrazolyl,dihydropyrrolyl, dihydropyrazinyl, dihydropyridinyl, dihydropyrimidinyl,dihydrofuryl, dihydropyranyl, tetrahydrothienyl S-oxide,tetrahydrothienyl S,S-dioxide and homothiomorpholinyl S-oxide. Theheterocycle groups herein are unsubstituted or, as specified,substituted in one or more substitutable positions with various groups.For example, such heterocycle groups may be optionally substituted withC₁-C₆ alkyl, C₁-C₆ alkoxy, halogen, hydroxy, cyano, nitro, amino,mono(C₁-C₆)alkylamino, di(C₁-C₆)alkylamino, C₂-C₆alkenyl, C₂-C₆alkynyl,C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, amino(C₁-C₆)alkyl,mono(C₁-C₆)alkylamino(C₁-C₆) alkyl, di(C₁-C₆)alkylamino(C₁-C₆) alkyl or═O.

Synthesis

The present invention provides the compounds (IX) for treating andpreventing Alzheimer's disease. The anti-Alzheimer's compounds (IX) aremade by methods well known to those skilled in the art from startingcompounds known to those skilled in the art. The process chemistry iswell known to those skilled in the art. The most general process toprepare the compounds (IX) of the present invention is set forth inCHART A. The chemistry is straight forward and in summary involves thesteps of alkylating a hydroxyl substituted amino acid starting material(I) to produce the corresponding N-protected amino acid (II). Reactionof the N-protected amino acid (II) with diazomethane (where R₂ and R₃are H) followed by a reduction produces the corresponding alcoholcompound (III). Subsequent formation of the corresponding epoxide (IV),followed by ring opening of the epoxide (V) with a C-terminal amine,R_(c)—NH₂ (V) produces the corresponding protected alcohol (VI). Thenitrogen protecting group of (VI) is removed to produce thecorresponding primary amine which is then reacted with a nitrogenprotected (for example Boc) amino acid of the formula Boc-NH—CH(Y)—CO₂Hto produce coupled amine (VII). The coupled amines (VII) are saponifiedand further nitrogen-deprotected to provide cyclization precursor (VIII)which is then cyclized to provide the anti-Alzheimer's compounds (IX).One skilled in the art will appreciate that these are all well knownreactions in organic chemistry. A chemist skilled in the art, knowingthe chemical structure of the biologically active compound end product(IX) of the invention would be able to prepare them by known methodsfrom known starting materials without any additional information. Theexplanation below therefore is not necessary but is deemed helpful tothose skilled in the art who desire to make the compounds of the presentinvention. Prefered methods include, but are not limited to, thosemethods described below.

The —NH—CH(R)—CH(OH)— moiety of the compounds of formula (IX) can bereadily prepared by methods disclosed in the literature and known tothose skilled in the art. For example, J. Med. Chem., 36, 288-291(1993), Tetrahedron Letters, 28, 5569-5572 (1987), J. Med. Chem., 38,581-584 (1995) and Tetrahedron Letters, 38, 619-620 (1997) all discloseprocesses to prepare hydroxyethylamine type compounds.

CHARTS A-C set forth a general method used in the present invention toprepare the appropriate compounds of formula (IX). The compounds offormula (IX) of the present invention are prepared by starting with thehydroxyl substituted amino acid starting materials (I). These are wellknown to those skilled in the art, are commercially available, or can bereadily prepared from known compounds by methods well known to thoseskilled in the art. The compounds of formula (IX) of the presentinvention have at least two or three enantiomeric centers. The presentinvention relates to all diastereomers and enantiomers.

The first step of the process is alkylation of the hydroxyl group of theN-protected amino acid ester (I). Further guidance may be found in J.Med. Chem. 43, 1271, (2000). It is prefered that the nitrogen protectinggroup be t-butoxycarbonyl (BOC) or benzyloxycarbonyl (CBZ), it is moreprefered that the protecting group be t-butoxycarbonyl. One skilled inthe art will understand the prefered methods of introducing at-butoxycarbonyl or benzyloxycarbonyl protecting group and mayadditionally consult T. W. Green and P. G. M. Wuts in “Protective Groupsin Organic Chemistry, John Wiley and Sons, 1991 for guidance.

The N-protected amino acid (II) is transformed to the correspondingN-protected compound (III) following a two step procedure. If it isdesired that both R₂ and R₃ are —H, then the N-protected amino acid (II)is reacted with diazomethane, as is well known to those skilled in theart. X₁ includes —Cl, —Br, —I, —O-tosylate, —O-mesylate, —O-nosylate.;it is prefered that —X₁ be —Br or —Cl. Suitable reaction conditionsinclude running the reaction in inert solvents, such as but not limitedto diethyl ether, tetrahydrofuran and the like. The reactions from theN-protected amino acid (II) are carried out for a period of time between10 minutes and 1 day and at temperatures ranging from −78° to 20-25°. Itis preffered to conduct the reactions for a period of time between 1-4hours and at temperatures between −30° to −10°. This process adds onemethylene group.

Alternatively, the compounds of formula (III) can be formed by firstconverting the N-protected amino acid (II) to a corresponding methyl orethyl ester, according to methods well established in the art, followedby treatment with a reagent of formula X₁—C(R₂)(R₃)—X₁ and a strongmetal base. The base serves to affect a halogen-metal exchange, wherethe —X₁ undergoing exchange is a halogen selected from chlorine, bromineor iodine. Suitable bases include, but are not limited to thealkyllithiums including, for example, sec-butyllithium, n-butyllithium,and t-butyllithium. The reactions are preferably conducted at lowtemperature, such as −78°. Suitable reaction conditions include runningthe reaction in inert solvents, such as but not limited to, ether,tetrahydrofuran and the like. Where R₂ and R₃ are both hydrogen, thenexamples of X₁—C(R₂)(R₃)—X₁ include dibromomethane, diiodomethane,chloroiodomethane, bromoiodomethane and bromochloromethane. One skilledin the art knows the prefered conditions required to conduct thisreaction. Furthermore, if R₂ and/or R₃ are not —H, then by the additionof —C(R₂)(R₃)—X₁ to esters of the N-protected amino acid (II), anadditional chiral center will be incorporated into the product, providedthat R₂ and R₃ are not the same.

After addition, the intermediate ketone is then reduced by means wellknown to those skilled in the art for reduction of a ketone to thecorresponding secondary alcohol affording the corresponding alcohol(III). The means and reaction conditions for reducing the intermediateketone to the corresponding alcohol (III) include, for example, sodiumborohydride, lithium borohydride, borane, diisobutylaluminum hydride,and lithium aluminium hydride. Sodium borohydride is a prefered reducingagent. The reductions are carried out for a period of time between 1hour and 3 days at temperatures ranging from −78° to elevatedtemperature up to the reflux point of the solvent employed. It isprefered to conduct the reduction between −78° and 0°. If borane isused, it may be employed as a complex, for example, borane-methylsulfide complex, borane-piperidine complex, or borane-tetrahydrofurancomplex. The prefered combination of reducing agents and reactionconditions needed are known to those skilled in the art, see forexample, Larock, R. C. in Comprehensive Organic Transformations, VCHPublishers, 1989. The conversion of the protected compound of formula(II) to the corresponding alcohol (III) produces the second chiralcenter (third chiral center if R₂ and R₃ are not the same). Thereduction produces a mixture of enantiomers at the second center ofalcohol (III). This diastereomeric and enantiomeric mixture may beseparated by means known to those skilled in the art such as selectivelow-temperature recrystallization or chromatographic separation, forexample by HPLC, employing commercially available chiral columns.

The alcohol (III) is transformed to the corresponding epoxide (IV) bymeans known to those skilled in the art. A prefered means is by reactionwith base, for example, but not limited to, hydroxide ion generated fromsodium hydroxide, potassium hydroxide, lithium hydroxide and the like.Reaction conditions include the use of C₁-C₆ alcohol solvents; ethanolis prefered. A common co-solvent, such as for example, ethyl acetate mayalso be employed. Reactions are conducted at temperatures ranging from−45° up to the reflux temperature of the alcohol employed; preferedtemperature ranges are between −20° and 20-25°.

The epoxide (IV) is then reacted with the appropriately substitutedC-terminal amine, R_(C)—NH₂ (V) by means known to those skilled in theart which opens the epoxide to produce the desired correspondingprotected alcohol (VI). The substituted C-terminal amines, R_(C)—NH₂ (V)of this invention are commercially available or are known to thoseskilled in the art and can be readily prepared from known compounds.Suitable reaction conditions for opening the epoxide (IV) includerunning the reaction in a wide range of common and inert solvents. C₁-C₆alcohol solvents are prefered and isopropyl alcohol most prefered. Thereactions can be run at temperatures ranging from 20-25° up to thereflux temperature of the alcohol employed. The prefered temperaturerange for conducting the reaction is between 50° up to the refluxtemperature of the alcohol employed. When the substituted C-terminalamine (V) is an aminomethyl group where the substituent on the methylgroup is an aryl group, for example NH₂—CH₂—R_(C-aryl), andNH₂—CH₂—R_(C-aryl) is not commercially available it is preferrablyprepared as follows. A suitable starting material is the (appropriatelysubstituted) aralkyl compound. The first step is bromination of thealkyl substitutent via methods known to those skilled in the art, seefor example R. C. Larock in Comprehensive Organic Transformations, VCHPublishers, 1989, p. 313. Next the alkyl halide is reacted with azide toproduce the aryl-(alkyl)-azide. Last, the azide is reduced to thecorresponding amine by hydrogen/catalyst to give the C-terminal amine(V) of formula NH₂—CH₂—R_(C-aryl).

The protected alcohol (VI) is nitrogen-deprotected by means known tothose skilled in the art for removal of amine protecting group. Suitablemeans for removal of the amine protecting group depends on the nature ofthe protecting group. Those skilled in the art, knowing the nature of aspecific protecting group, know which reagent is preferable for itsremoval. For example, it is prefered to remove the prefered protectinggroup, BOC, by dissolving the protected alcohol (VI) in atrifluoroacetic acid/dichloromethane (1/1) mixture. When complete, thesolvents are removed under reduced pressure to give the correspondingamine (as the corresponding salt, i.e. trifluoroacetic acid salt) whichis used without further purification. However, if desired, the amine canbe purified further by means well known to those skilled in the art,such as for example, recrystallization. Further, if the non-salt form isdesired that also can be obtained by, such as for example, preparing thefree base amine via treatment of the salt with mild basic conditions.Additional BOC deprotection conditions and deprotection conditions forother protecting groups can be found in T. W. Green and P. G. M. Wuts in“Protective Groups in Organic Chemistry, John Wiley and Sons, 1991, p.309. Suitable chemically suitable salts include trifluoroacetate, andthe anion of mineral acids such as chloride, sulfate, phosphate;prefered is trifluoroacetate.

The deprotected amine is then reacted with an appropriately substitutedamide forming agent of the formula Boc-NH—CH(Y)—CO₂H to produce coupledamines (VII) by nitrogen-acylation means known to those skilled in theart. Nitrogen acylation conditions for reaction of the deprotected aminewith an amide forming agent to produce the corresponding coupled amine(VII) are known to those skilled in the art and can be found in R. C.Larock in Comprehensive Organic Transformations, VCH Publishers, 1989,p. 981, 979, and 972. The nitrogen-acylation of primary amines toproduce secondary amides is one of the oldest known reactions. The amideforming agents of the formula Boc-NH—CH(Y)—CO₂H are readily preparedfrom known starting materials by methods known in the literature. Forfurther guidance may be found in Bodanszky, M. “Principles of PeptideSynthesis, 2^(nd) Edition, Springer Verlag, 1993.

The coupled amines (VII) are further deprotected as illustrated in CHARTA to afford cyclization precursors (VIII). Numerous conditions exist fordeprotection and are understood by those skilled in the art;alternatively, one may consult T. W. Green and P. G. M. Wuts in“Protective Groups in Organic Chemistry, John Wiley and Sons, 1991, p.49. The cyclization precursors are then exposed to standardmacrolactamization chemistry to provide the title compounds (IX) ;conditions for effecting this reaction with correspondingmacrocyclization are amply documented in the primary literature.

CHART B sets forth an alternative route to compounds of formula (IX) fortreating and preventing Alzhiemer's disease. The compounds of formula(IX) are made by methods well known to those skilled in the art fromstarting materials known to those skilled in the art. The processchemistry is well known to those skilled in the art. The chemistry isstraight forward and follows many of the generalizations described forCHART A. In CHART B X₄ is defined as an oxygen protecting group that canbe removed simultaneously with the nitrogen protecting group of (XIV),for example benzyl. One skilled in the art will understand the preferredmethods of removing such groups and may additionally consult T. W. Greenand P. G. M. Wuts in “Protective Groups in Organic Syntheis”, John Wileyand Sons, 1999 for guidance. The protected amino acids of the formula(XIV) are commercially available or readily prepared by methods known inthe literature. Z comprises —OH (carboxylic acid) or halide (acylhalide), preferably chlorine, or a suitable group to produce a mixedanhydride. The diol of (XIII) is transformed to the correspondingepoxide by means known to those skilled in the art. A preferred means isby reaction with diisopropylazodicarboxylate in the presence oftriphenylphosphine. Additionally one can consult Tetrahedron 1992, 48,10515 and references therein for further guidance. The amide formingagents of the formula L-X—CO-Z (XVIII) are commercially available orreadily prepared by methods known in the literature. Z comprises —OH(carboxylic acid) or halide (acyl halide), preferably chlorine, or asuitable group to produce a mixed anhydride. L comprises halide,preferably bromine or iodine or OH, or a complimentary functionalitythat will result in bond formation with the OH substituent of B. CHART Bis further exemplified by the synthesis of12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione(1) in Example one.

CHART C sets forth an alternative route to compounds (IX) for treatingand preventing Alzhiemer's disease. The compounds of formula (IX) aremade by methods well known to those skilled in the art from startingmaterials known to those skilled in the art. The process chemistry iswell known to those skilled in the art. The chemistry is straightforward and follows many of the generalizations described for CHART A.In CHART C X₄ is defined as an oxygen protecting group that can beremoved simultaneously with the nitrogen protecting group of (XX), forexample benzyl. One skilled in the art will understand the preferredmethods of removing such groups and may additionally consult T. W. Greenand P. G. M. Wuts in “Protective Groups in Organic Syntheis”, John Wileyand Sons, 1999 for guidance. The protected amino acids of the formula(XIV) are commercially available or readily prepared by methods known inthe literature. Z comprises —OH (carboxylic acid) or halide (acylhalide), preferably chlorine, or a suitable group to produce a mixedanhydride. The amide forming agents of the formula L-X—CO-Z (XVIII) arecommercially available or readily prepared by methods known in theliterature. Z comprises —OH (carboxylic acid) or halide (acyl halide),preferably chlorine, or a suitable group to produce a mixed anhydride. Lcomprises halide, preferably bromine or iodine or OH, or a complimentaryfunctionality that will result in bond formation with the OH substituentof B. One skilled in the art will understand the preferred methods ofintroducing and removing cyclic carbonate protecting groups and mayadditionally consult T. W. Green and P. G. M. Wuts in “Protective Groupsin Organic Syntheis”, John Wiley and Sons, 1999 for guidance. The diol(XXIII) is transformed to the corresponding epoxide by means known tothose skilled in the art. A preferred means is by reaction with1-(p-toluenesulfonyl) imidazole followed by potassium t-butoxide. SeeTetrahedron Asymmetry, 1999, 10, 837. Additionally one can consultTetrahedron 1992, 48, 10515 and references therein for further guidance.

Methods of the Invention

The compounds of the invention, and pharmaceutically acceptable saltsthereof, are suitable for treating humans and/or animals suffering froma condition characterized by a pathological form of beta-amyloidpeptide, such as beta-amyloid plaques, and for helping to prevent ordelay the onset of such a condition. For example, the compounds are fortreating Alzheimer's disease, for helping prevent or delay the onset ofAlzheimer's disease, for treating patients with MCI (mild cognitiveimpairment) and preventing or delaying the onset of Alzheimer's diseasein those who would progress from MCI to AD, for treating Down'ssyndrome, for treating humans who have Hereditary Cerebral Hemorrhagewith Amyloidosis of the Dutch-Type, for treating cerebral amyloidangiopathy and preventing its potential consequences, i.e. single andrecurrent lobal hemorrhages, for treating other degenerative dementias,including dementias of mixed vascular and degenerative origin, dementiaassociated with Parkinson's disease, dementia associated withprogressive supranuclear palsy, dementia associated with cortical basaldegeneration, and diffuse Lewy body type Alzheimer's disease. Thecompounds and compositions of the invention are particularly suitablefor treating or preventing Alzheimer's disease. When treating orpreventing these diseases, the compounds of the invention can either beused individually or in combination, as is best for the patient.

As used herein, the term “treating” means that the compounds of theinvention can be used in humans with at least a tentative diagnosis ofdisease. The compounds of the invention will delay or slow theprogression of the disease thereby giving the individual a more usefullife span.

The term “preventing” means that the compounds of the present inventionare administered to a patient who has not been diagnosed as possiblyhaving the disease at the time of administration, but who would normallybe expected to develop the disease or be at increased risk for thedisease. The compounds of the invention will slow the development ofdisease symptoms, delay the onset of the disease, or prevent theindividual from developing the disease at all. Preventing also includesadministration of the compounds of the invention to those individualsthought to be predisposed to the disease due to age, familial history,genetic or chromosomal abnormalities, and/or due to the presence of oneor more biological markers for the disease, such as a known geneticmutation of APP or APP cleavage products in brain tissues or fluids.

In treating or preventing the above diseases, the compounds of theinvention are administered in a therapeutically effective amount. Thetherapeutically effective amount will vary depending on the particularcompound used and the route of administration, as is known to thoseskilled in the art.

In treating a patient displaying any of the diagnosed above conditions aphysician may administer a compound of the invention immediately andcontinue administration indefinitely, as needed. In treating patientswho are not diagnosed as having Alzheimer's disease, but who arebelieved to be at substantial risk for Alzheimer's disease, thephysician should preferably start treatment when the patient firstexperiences early pre-Alzheimer's symptoms such as, memory or cognitiveproblems associated with aging. In addition, there are some patients whomay be determined to be at risk for developing Alzheimer's through thedetection of a genetic marker such as APOE4 or other biologicalindicators that are predictive for Alzheimer's disease. In thesesituations, even though the patient does not have symptoms of thedisease, administration of the compounds of the invention may be startedbefore symptoms appear, and treatment may be continued indefinitely toprevent or delay the outset of the disease.

Dosage Forms and Amounts

The compounds of the invention can be administered orally,parenternally, (IV, IM, depo-IM, SQ, and depo SQ), sublingually,intranasally (inhalation), intrathecally, topically, or rectally. Dosageforms known to those of skill in the art are suitable for delivery ofthe compounds of the invention.

Compositions are provided that contain therapeutically effective amountsof the compounds of the invention. The compounds are preferablyformulated into suitable pharmaceutical preparations such as tablets,capsules, or elixirs for oral administration or in sterile solutions orsuspensions for parenternal administration. Typically the compoundsdescribed above are formulated into pharmaceutical compositions usingtechniques and procedures well known in the art.

About 1 to 500 mg of a compound or mixture of compounds of the inventionor a physiologically acceptable salt or ester is compounded with aphysiologically acceptable vehicle, carrier, excipient, binder,preservative, stabilizer, flavor, etc., in a unit dosage form as calledfor by accepted pharmaceutical practice. The amount of active substancein those compositions or preparations is such that a suitable dosage inthe range indicated is obtained. The compositions are preferablyformulated in a unit dosage form, each dosage containing from about 2 toabout 100 mg, more preferably about 10 to about 30 mg of the activeingredient. The term “unit dosage from” refers to physically discreteunits suitable as unitary dosages for human subjects and other mammals,each unit containing a predetermined quantity of active materialcalculated to produce the desired therapeutic effect, in associationwith a suitable pharmaceutical excipient.

To prepare compositions, one or more compounds of the invention aremixed with a suitable pharmaceutically acceptable carrier. Upon mixingor addition of the compound(s), the resulting mixture may be a solution,suspension, emulsion, or the like. Liposomal suspensions may also besuitable as pharmaceutically acceptable carriers. These may be preparedaccording to methods known to those skilled in the art. The form of theresulting mixture depends upon a number of factors, including theintended mode of administration and the solubility of the compound inthe selected carrier or vehicle. The effective concentration issufficient for lessening or ameliorating at least one symptom of thedisease, disorder, or condition treated and may be empiricallydetermined.

Pharmaceutical carriers or vehicles suitable for administration of thecompounds provided herein include any such carriers known to thoseskilled in the art to be suitable for the particular mode ofadministration. In addition, the active materials can also be mixed withother active materials that do not impair the desired action, or withmaterials that supplement the desired action, or have another action.The compounds may be formulated as the sole pharmaceutically activeingredient in the composition or may be combined with other activeingredients.

Where the compounds exhibit insufficient solubility, methods forsolubilizing may be used. Such methods are known and include, but arenot limited to, using cosolvents such as dimethylsulfoxide (DMSO), usingsurfactants such as Tween®, and dissolution in aqueous sodiumbicarbonate. Derivatives of the compounds, such as salts or prodrugs mayalso be used in formulating effective pharmaceutical compositions.

The concentration of the compound is effective for delivery of an amountupon administration that lessens or ameliorates at least one symptom ofthe disorder for which the compound is administered. Typically, thecompositions are formulated for single dosage administration.

The compounds of the invention may be prepared with carriers thatprotect them against rapid elimination from the body, such astime-release formulations or coatings. Such carriers include controlledrelease formulations, such as, but not limited to, microencapsulateddelivery systems. The active compound is included in thepharmaceutically acceptable carrier in an amount sufficient to exert atherapeutic effect in the absence of undesirable side effects on thepatient treated. The therapeutically effective concentration may bedetermined empirically by testing the compounds in known in vitro and invivo model systems for the treated disorder.

The compounds and compositions of the invention can be enclosed inmultiple or single dose containers. The enclosed compounds andcompositions can be provided in kits, for example, including componentparts that can be assembled for use. For example, a compound inhibitorin lyophilized form and a suitable diluent may be provided as separatedcomponents for combination prior to use. A kit may include a compoundinhibitor and a second therapeutic agent for co-administration. Theinhibitor and second therapeutic agent may be provided as separatecomponent parts. A kit may include a plurality of containers, eachcontainer holding one or more unit dose of the compound of theinvention. The containers are preferably adapted for the desired mode ofadministration, including, but not limited to tablets, gel capsules,sustained-release capsules, and the like for oral administration; depotproducts, pre-filled syringes, ampules, vials, and the like forparenternal administration; and patches, medipads, creams, and the likefor topical administration.

The concentration of active compound in the drug composition will dependon absorption, inactivation, and excretion rates of the active compound,the dosage schedule, and amount administered as well as other factorsknown to those of skill in the art.

The active ingredient may be administered at once, or may be dividedinto a number of smaller doses to be administered at intervals of time.It is understood that the precise dosage and duration of treatment is afunction of the disease being treated and may be determined empiricallyusing known testing protocols or by extrapolation from in vivo or invitro test data. It is to be noted that concentrations and dosage valuesmay also vary with the severity of the condition to be alleviated. It isto be further understood that for any particular subject, specificdosage regimens should be adjusted over time according to the individualneed and the professional judgment of the person administering orsupervising the administration of the compositions, and that theconcentration ranges set forth herein are exemplary only and are notintended to limit the scope or practice of the claimed compositions.

If oral administration is desired, the compound should be provided in acomposition that protects it from the acidic environment of the stomach.For example, the composition can be formulated in an enteric coatingthat maintains its integrity in the stomach and releases the activecompound in the intestine. The composition may also be formulated incombination with an antacid or other such ingredient.

Oral compositions will generally include an inert diluent or an ediblecarrier and may be compressed into tablets or enclosed in gelatincapsules. For the purpose of oral therapeutic administration, the activecompound or compounds can be incorporated with excipients and used inthe form of tablets, capsules, or troches. Pharmaceutically compatiblebinding agents and adjuvant materials can be included as part of thecomposition.

The tablets, pills, capsules, troches, and the like can contain any ofthe following ingredients or compounds of a similar nature: a bindersuch as, but not limited to, gum tragacanth, acacia, corn starch, orgelatin; an excipient such as microcrystalline cellulose, starch, orlactose; a disintegrating agent such as, but not limited to, alginicacid and corn starch; a lubricant such as, but not limited to, magnesiumstearate; a gildant, such as, but not limited to, colloidal silicondioxide; a sweetening agent such as sucrose or saccharin; and aflavoring agent such as peppermint, methyl salicylate, or fruitflavoring.

When the dosage unit form is a capsule, it can contain, in addition tomaterial of the above type, a liquid carrier such as a fatty oil. Inaddition, dosage unit forms can contain various other materials, whichmodify the physical form of the dosage unit, for example, coatings ofsugar and other enteric agents. The compounds can also be administeredas a component of an elixir, suspension, syrup, wafer, chewing gum orthe like. A syrup may contain, in addition to the active compounds,sucrose as a sweetening agent and certain preservatives, dyes andcolorings, and flavors.

The active materials can also be mixed with other active materials thatdo not impair the desired action, or with materials that supplement thedesired action.

Solutions or suspensions used for parenternal, intradermal,subcutaneous, or topical application can include any of the followingcomponents: a sterile diluent such as water for injection, salinesolution, fixed oil, a naturally occurring vegetable oil such as sesameoil, coconut oil, peanut oil, cottonseed oil, and the like, or asynthetic fatty vehicle such as ethyl oleate, and the like, polyethyleneglycol, glycerine, propylene glycol, or other synthetic solvent;antimicrobial agents such as benzyl alcohol and methyl parabens;antioxidants such as ascorbic acid and sodium bisulfite; chelatingagents such as ethylenediaminetetraacetic acid (EDTA); buffers such asacetates, citrates, and phosphates; and agents for the adjustment oftonicity such as sodium chloride and dextrose. Parenternal preparationscan be enclosed in ampoules, disposable syringes, or multiple dose vialsmade of glass, plastic, or other suitable material. Buffers,preservatives, antioxidants, and the like can be incorporated asrequired.

Where administered intravenously, suitable carriers includephysiological saline, phosphate buffered saline (PBS), and solutionscontaining thickening and solubilizing agents such as glucose,polyethylene glycol, polypropyleneglycol, and mixtures thereof.Liposomal suspensions including tissue-targeted liposomes may also besuitable as pharmaceutically acceptable carriers. These may be preparedaccording to methods known for example, as described in U.S. Pat. No.4,522,811.

The active compounds may be prepared with carriers that protect thecompound against rapid elimination from the body, such as time-releaseformulations or coatings. Such carriers include controlled releaseformulations, such as, but not limited to, implants andmicroencapsulated delivery systems, and biodegradable, biocompatiblepolymers such as collagen, ethylene vinyl acetate, polyanhydrides,polyglycolic acid, polyorthoesters, polylactic acid, and the like.Methods for preparation of such formulations are known to those skilledin the art.

The compounds of the invention can be administered orally, parenternally(IV, IM, depo-IM, SQ, and depo-SQ), sublingually, intranasally(inhalation), intrathecally, topically, or rectally. Dosage forms knownto those skilled in the art are suitable for delivery of the compoundsof the invention.

Compounds of the invention may be administered enterally orparenterally. When administered orally, compounds of the invention canbe administered in usual dosage forms for oral administration as is wellknown to those skilled in the art. These dosage forms include the usualsolid unit dosage forms of tablets and capsules as well as liquid dosageforms such as solutions, suspensions, and elixirs. When the solid dosageforms are used, it is preferred that they be of the sustained releasetype so that the compounds of the invention need to be administered onlyonce or twice daily.

The oral dosage forms are administered to the patient 1, 2, 3, or 4times daily. It is preferred that the compounds of the invention beadministered either three or fewer times, more preferably once or twicedaily. Hence, it is preferred that the compounds of the invention beadministered in oral dosage form. It is preferred that whatever oraldosage form is used, that it be designed so as to protect the compoundsof the invention from the acidic environment of the stomach. Entericcoated tablets are well known to those skilled in the art. In addition,capsules filled with small spheres each coated to protect from theacidic stomach, are also well known to those skilled in the art.

When administered orally, an administered amount therapeuticallyeffective to inhibit beta-secretase activity, to inhibit A betaproduction, to inhibit A beta deposition, or to treat or prevent AD isfrom about 0.1 mg/day to about 1,000 mg/day. It is preferred that theoral dosage is from about 1 mg/day to about 100 mg/day. It is morepreferred that the oral dosage is from about 5 mg/day to about 50mg/day. It is understood that while a patient may be started at onedose, that dose may be varied over time as the patient's conditionchanges.

Compounds of the invention may also be advantageously delivered in anano crystal dispersion formulation. Preparation of such formulations isdescribed, for example, in U.S. Pat. No. 5,145,684. Nano crystallinedispersions of HIV protease inhibitors and their method of use aredescribed in U.S. Pat. No. 6,045,829. The nano crystalline formulationstypically afford greater bioavailability of drug compounds.

The compounds of the invention can be administered parenterally, forexample, by IV, IM, depo-IM, SC, or depo-SC. When administeredparenterally, a therapeutically effective amount of about 0.5 to about100 mg/day, preferably from about 5 to about 50 mg daily should bedelivered. When a depot formulation is used for injection once a monthor once every two weeks, the dose should be about 0.5 mg/day to about 50mg/day, or a monthly dose of from about 15 mg to about 1,500 mg. In partbecause of the forgetfulness of the patients with Alzheimer's disease,it is preferred that the parenteral dosage form be a depo formulation.

The compounds of the invention can be administered sublingually. Whengiven sublingually, the compounds of the invention should be given oneto four times daily in the amounts described above for IMadministration.

The compounds of the invention can be administered intranasally. Whengiven by this route, the appropriate dosage forms are a nasal spray ordry powder, as is known to those skilled in the art. The dosage of thecompounds of the invention for intranasal administration is the amountdescribed above for IM administration.

The compounds of the invention can be administered intrathecally. Whengiven by this route the appropriate dosage form can be a parenternaldosage form as is known to those skilled in the art. The dosage of thecompounds of the invention for intrathecal administration is the amountdescribed above for IM administration.

The compounds of the invention can be administered topically. When givenby this route, the appropriate dosage form is a cream, ointment, orpatch. Because of the amount of the compounds of the invention to beadministered, the patch is preferred. When administered topically, thedosage is from about 0.5 mg/day to about 200 mg/day. Because the amountthat can be delivered by a patch is limited, two or more patches may beused. The number and size of the patch is not important, what isimportant is that a therapeutically effective amount of the compounds ofthe invention be delivered as is known to those skilled in the art. Thecompounds of the invention can be administered rectally by suppositoryas is known to those skilled in the art. When administered bysuppository, the therapeutically effective amount is from about 0.5 mgto about 500 mg.

The compounds of the invention can be administered by implants as isknown to those skilled in the art. When administering a compound of theinvention by implant, the therapeutically effective amount is the amountdescribed above for depot administration.

The invention here is the new compounds of the invention and new methodsof using the compounds of the invention. Given a particular compound ofthe invention and a desired dosage form, one skilled in the art wouldknow how to prepare and administer the appropriate dosage form.

The compounds of the invention are used in the same manner, by the sameroutes of administration, using the same pharmaceutical dosage forms,and at the same dosing schedule as described above, for preventingdisease or treating patients with MCI (mild cognitive impairment) andpreventing or delaying the onset of Alzheimer's disease in those whowould progress from MCI to AD, for treating or preventing Down'ssyndrome, for treating humans who have Hereditary Cerebral Hemorrhagewith Amyloidosis of the Dutch-Type, for treating cerebral amyloidangiopathy and preventing its potential consequences, i.e. single andrecurrent lobar hemorrhages, for treating other degenerative dementias,including dementias of mixed vascular and degenerative origin, dementiaassociated with Parkinson's disease, dementia associated withprogressive supranuclear palsy, dementia associated with cortical basaldegeneration, and diffuse Lewy body type of Alzheimer's disease.

The compounds of the invention can be used in combination, with eachother or with other therapeutic agents or approaches used to treat orprevent the conditions listed above. Such agents or approaches include:acetylcholine esterase inhibitors such as tacrine(tetrahydroaminoacridine, marketed as COGNEX®), donepezil hydrochloride,(marketed as Aricept® and rivastigmine (marketed as Exelon®) ;gamma-secretase inhibitors; anti-inflammatory agents such ascyclooxygenase II inhibitors; anti-oxidants such as Vitamin E andginkolides; immunological approaches, such as, for example, immunizationwith A beta peptide or administration of anti-A beta peptide antibodies;statins; and direct or indirect neurotropic agents such asCerebrolysin®, AIT-082 (Emilieu, 2000, Arch. Neurol. 57:454), and otherneurotropic agents of the future.

In addition, the compounds of the present invention can also be usedwith inhibitors of P-glycoproten (P-gp). The use of P-gp inhibitors isknown to those skilled in the art. See for example, Cancer Research, 53,4595-4602 (1993), Clin. Cancer Res., 2, 7-12 (1996), Cancer Research,56, 4171-4179 (1996), International Publications WO99/64001 andWO01/10387. The important thing is that the blood level of the P-gpinhibitor be such that it exerts its effect in inhibiting P-gp fromdecreasing brain blood levels of the compounds of the present invention.To that end the P-gp inhibitor and the compounds of the presentinvention can be administered at the same time, by the same or differentroute of administration, or at different times. The important thing isnot the time of administration but having an effective blood level ofthe P-gp inhibitor.

Suitable P-gp inhibitors include cyclosporin A, verapamil, tamoxifen,quinidine, Vitamin E-TGPS, ritonavir, megestrol acetate, progesterone,rapamycin, 10,11-methanodibenzosuberane, phenothiazines, acridinederivatives such as GF120918, FK506, VX-710, LY335979, PSC-833,GF-102,918 and other steroids. It is to be understood that additionalagents will be found that do the same function.

The P-gp inhibitors can be administered orally, parenterally, (IV, IM,IM-depo, SQ, SQ-depo), topically, sublingually, rectally, intranasally,intrathecally and by implant.

The therapeutically effective amount of the P-gp inhibitors is fromabout 0.1 to about 300 mg/kg/day, preferably about 0.1 to about 150mg/kg daily. It is understood that while a patient may be started on onedose, that dose may have to be varied over time as the patient'scondition changes.

When administered orally, the P-gp inhibitors can be administered inusual dosage forms for oral administration as is known to those skilledin the art. These dosage forms include the usual solid unit dosage formsof tablets and capsules as well as liquid dosage forms such assolutions, suspensions and elixirs. When the solid dosage forms areused, it is preferred that they be of the sustained release type so thatthe P-gp inhibitors need to be administered only once or twice daily.The oral dosage forms are administered to the patient one thru fourtimes daily. It is preferred that the P-gp inhibitors be administeredeither three or fewer times a day, more preferably once or twice daily.Hence, it is preferred that the P-gp inhibitors be administered in soliddosage form and further it is preferred that the solid dosage form be asustained release form which permits once or twice daily dosing. It ispreferred that what ever dosage form is used, that it be designed so asto protect the P-gp inhibitors from the acidic environment of thestomach. Enteric coated tablets are well known to those skilled in theart. In addition, capsules filled with small spheres each coated toprotect from the acidic stomach, are also well known to those skilled inthe art.

In addition, the P-gp inhibitors can be administered parenterally. Whenadministered parenterally they can be administered IV, IM, depo-IM, SQor depo-SQ.

The P-gp inhibitors can be given sublingually. When given sublingually,the P-gp inhibitors should be given one thru four times daily in thesame amount as for IM administration.

The P-gp inhibitors can be given intranasally. When given by this routeof administration, the appropriate dosage forms are a nasal spray or drypowder as is known to those skilled in the art. The dosage of the P-gpinhibitors for intranasal administration is the same as for IMadministration.

The P-gp inhibitors can be given intrathecally. When given by this routeof administration the appropriate dosage form can be a parenteral dosageform as is known to those skilled in the art.

The P-gp inhibitors can be given topically. When given by this route ofadministration, the appropriate dosage form is a cream, ointment orpatch. Because of the amount of the P-gp inhibitors needed to beadministered the patch is preferred. However, the amount that can bedelivered by a patch is limited. Therefore, two or more patches may berequired. The number and size of the patch is not important, what isimportant is that a therapeutically effective amount of the P-gpinhibitors be delivered as is known to those skilled in the art.

The P-gp inhibitors can be administered rectally by suppository as isknown to those skilled in the art.

The P-gp inhibitors can be administered by implants as is known to thoseskilled in the art.

There is nothing novel about the route of administration or the dosageforms for administering the P-gp inhibitors. Given a particular P-gpinhibitor, and a desired dosage form, one skilled in the art would knowhow to prepare the appropriate dosage form for the P-gp inhibitor.

It should be apparent to one skilled in the art that the exact dosageand frequency of administration will depend on the particular compoundsof the invention administered, the particular condition being treated,the severity of the condition being treated, the age, weight, generalphysical condition of the particular patient, and other medication theindividual may be taking as is well known to administering physicianswho are skilled in this art.

Inhibition of APP Cleavage

The compounds of the invention inhibit cleavage of APP between Met595and Asp596 numbered for the APP695 isoform, or a mutant thereof, or at acorresponding site of a different isoform, such as APP751 or APP770, ora mutant thereof (sometimes referred to as the “beta secretase site”).While not wishing to be bound by a particular theory, inhibition ofbeta-secretase activity is thought to inhibit production of beta amyloidpeptide (A beta). Inhibitory activity is demonstrated in one of avariety of inhibition assays, whereby cleavage of an APP substrate inthe presence of a beta-secretase enzyme is analyzed in the presence ofthe inhibitory compound, under conditions normally sufficient to resultin cleavage at the beta-secretase cleavage site. Reduction of APPcleavage at the beta-secretase cleavage site compared with an untreatedor inactive control is correlated with inhibitory activity. Assaysystems that can be used to demonstrate efficacy of the compoundinhibitors of the invention are known. Representative assay systems aredescribed, for example, in U.S. Pat. Nos. 5,942,400, 5,744,346, as wellas in the Examples below.

The enzymatic activity of beta-secretase and the production of A betacan be analyzed in vitro or in vivo, using natural, mutated, and/orsynthetic APP substrates, natural, mutated, and/or synthetic enzyme, andthe test compound. The analysis may involve primary or secondary cellsexpressing native, mutant, and/or synthetic APP and enzyme, animalmodels expressing native APP and enzyme, or may utilize transgenicanimal models expressing the substrate and enzyme. Detection ofenzymatic activity can be by analysis of one or more of the cleavageproducts, for example, by immunoassay, flurometric or chromogenic assay,HPLC, or other means of detection. Inhibitory compounds are determinedas those having the ability to decrease the amount of beta-secretasecleavage product produced in comparison to a control, wherebeta-secretase mediated cleavage in the reaction system is observed andmeasured in the absence of inhibitory compounds.

Beta-Secretase

Various forms of beta-secretase enzyme are known, and are available forassay of enzyme activity and inhibition of enzyme activity. Theseinclude native, recombinant, and synthetic forms of the enzyme. Humanbeta-secretase is known as Beta Site APP Cleaving Enzyme (BACE), Asp2,and memapsin 2, and has been characterized, for example, in U.S. Pat.No. 5,744,346 and published PCT patent applications WO98/22597,WO00/03819, WO01/23533, and WO00/17369, as well as in literaturepublications (Hussain et.al., 1999, Mol.Cell.Neurosci. 14:419-427;Vassar et.al., 1999, Science 286:735-741; Yan et.al., 1999, Nature402:533-537; Sinha et.al., 1999, Nature 40:537-540; and Lin et.al.,2000, PNAS USA 97:1456-1460). Synthetic forms of the enzyme have alsobeen described (WO98/22597 and WO00/17369). Beta-secretase can beextracted and purified from human brain tissue and can be produced incells, for example mammalian cells expressing recombinant enzyme.

Preferred compounds are effective to inhibit 50% of beta-secretaseenzymatic activity at a concentration of less than 50 micromolar,preferably at a concentration of 10 micromolar or less, more preferably1 micromolar or less, and most preferably 10 nanomolar or less.

APP Substrate

Assays that demonstrate inhibition of beta-secretase-mediated cleavageof APP can utilize any of the known forms of APP, including the 695amino acid “normal” isotype described by Kang et.al., 1987, Nature325:733-6, the 770 amino acid isotype described by Kitaguchi et. al.,1981, Nature 331:530-532, and variants such as the Swedish Mutation(KM670-1NL) (APP-SW), the London Mutation (V7176F), and others. See, forexample, U.S. Pat. No. 5,766,846 and also Hardy, 1992, Nature Genet.1:233-234, for a review of known variant mutations. Additionalsubstrates include the dibasic amino acid modification, APP-KKdisclosed, for example, in WO 00/17369, fragments of APP, and syntheticpeptides containing the beta-secretase cleavage site, wild type (WT) ormutated form, e.g., SW, as described, for example, in U.S. Pat. No5,942,400 and WO00/03819.

The APP substrate contains the beta-secretase cleavage site of APP(KM-DA or NL-DA) for example, a complete APP peptide or variant, an APPfragment, a recombinant or synthetic APP, or a fusion peptide.Preferably, the fusion peptide includes the beta-secretase cleavage sitefused to a peptide having a moiety useful for enzymatic assay, forexample, having isolation and/or detection properties. Such moietiesinclude, for example, an antigenic epitope for antibody binding, a labelor other detection moiety, a binding substrate, and the like.

Antibodies

Products characteristic of APP cleavage can be measured by immunoassayusing various antibodies, as described, for example, in Pirttila et.al.,1999, Neuro. Lett. 249:21-4, and in U.S. Pat. No. 5,612,486. Antibodiesused to detect A beta include, for example, the monoclonal antibody 6E10(Senetek, St. Louis, Mo.) that specifically recognizes an epitope onamino acids 1-16 of the A beta peptide; antibodies 162 and 164 (New YorkState Institute for Basic Research, Staten Island, N.Y.) that arespecific for human A beta 1-40 and 1-42, respectively; and antibodiesthat recognize the junction region of beta-amyloid peptide, the sitebetween residues 16 and 17, as described in U.S. Pat. No. 5,593,846.Antibodies raised against a synthetic peptide of residues 591 to 596 ofAPP and SW192 antibody raised against 590-596 of the Swedish mutationare also useful in immunoassay of APP and its cleavage products, asdescribed in U.S. Pat. Nos. 5,604,102 and 5,721,130.

Assay Systems

Assays for determining APP cleavage at the beta-secretase cleavage siteare well known in the art. Exemplary assays, are described, for example,in U.S. Pat. Nos. 5,744,346 and 5,942,400, and described in the Examplesbelow.

Cell Free Assays

Exemplary assays that can be used to demonstrate the inhibitory activityof the compounds of the invention are described, for example, inWO00/17369, WO 00/03819, and U.S. Pat. Nos. 5,942,400 and 5,744,346.Such assays can be performed in cell-free incubations or in cellularincubations using cells expressing a beta-secretase and an APP substratehaving a beta-secretase cleavage site.

An APP substrate containing the beat-secretase cleavage site of APP, forexample, a complete APP or variant, an APP fragment, or a recombinant orsynthetic APP substrate containing the amino acid sequence: KM-DA orNL-DA, is incubated in the presence of beta-secretase enzyme, a fragmentthereof, or a synthetic or recombinant polypeptide variant havingbeta-secretase activity and effective to cleave the beta-secretasecleavage site of APP, under incubation conditions suitable for thecleavage activity of the enzyme. Suitable substrates optionally includederivatives that may be fusion proteins or peptides that contain thesubstrate peptide and a modification to facilitate the purification ordetection of the peptide or its beta-secretase cleavage products.Modifications include the insertion of a known antigenic epitope forantibody binding; the linking of a label or detectable moiety, thelinking of a binding substrate, and the like.

Suitable incubation conditions for a cell-free in vitro assay include,for example: approximately 200 nanomolar to 10 micromolar substrate,approximately 10 to 200 picomolar enzyme, and approximately 0.1nanomolar to 10 micromolar inhibitor compound, in aqueous solution, atan approximate pH of 4-7, at approximately 37 degrees C., for a timeperiod of approximately 10 minutes to 3 hours. These incubationconditions are exemplary only, and can be varied as required for theparticular assay components and/or desired measurement system.Optimization of the incubation conditions for the particular assaycomponents should account for the specific beta-secretase enzyme usedand its pH optimum, any additional enzymes and/or markers that might beused in the assay, and the like. Such optimization is routine and willnot require undue experimentation.

One assay utilizes a fusion peptide having maltose binding protein (MBP)fused to the C-terminal 125 amino acids of APP-SW. The MBP portion iscaptured on an assay substrate by anti-MBP capture antibody. Incubationof the captured fusion protein in the presence of beta-secretase resultsin cleavage of the substrate at the beta-secretase cleavage site.Analysis of the cleavage activity can be, for example, by immunoassay ofcleavage products. One such immunoassay detects a unique epitope exposedat the carboxy terminus of the cleaved fusion protein, for example,using the antibody SW192. This assay is described, for example, in U.S.Pat. No 5,942,400.

Cellular Assay

Numerous cell-based assays can be used to analyze beta-secretaseactivity and/or processing of APP to release A beta. Contact of an APPsubstrate with a beta-secretase enzyme within the cell and in thepresence or absence of a compound inhibitor of the invention can be usedto demonstrate beta-secretase inhibitory activity of the compound.Preferably, assay in the presence of an inhibitory compound provides atleast about 30%, most preferably at least about 50% inhibition of theenzymatic activity, as compared with a non-inhibited control.

In one embodiment, cells that naturally express beta-secretase are used.Alternatively, cells are modified to express a recombinantbeta-secretase or synthetic variant enzyme as discussed above. The APPsubstrate may be added to the culture medium and is preferably expressedin the cells. Cells that naturally express APP, variant or mutant formsof APP, or cells transformed to express an isoform of APP, mutant orvariant APP, recombinant or synthetic APP, APP fragment, or syntheticAPP peptide or fusion protein containing the beta-secretase APP cleavagesite can be used, provided that the expressed APP is permitted tocontact the enzyme and enzymatic cleavage activity can be analyzed.

Human cell lines that normally process A beta from APP provide a meansto assay inhibitory activities of the compounds of the invention.Production and release of A beta and/or other cleavage products into theculture medium can be measured, for example by immunoassay, such asWestern blot or enzyme-linked immunoassay (EIA) such as by ELISA.

Cells expressing an APP substrate and an active beta-secretase can beincubated in the presence of a compound inhibitor to demonstrateinhibition of enzymatic activity as compared with a control. Activity ofbeta-secretase can be measured by analysis of one or more cleavageproducts of the APP substrate. For example, inhibition of beta-secretaseactivity against the substrate APP would be expected to decrease releaseof specific beta-secretase induced APP cleavage products such as A beta.

Although both neural and non-neural cells process and release A beta,levels of endogenous beta-secretase activity are low and often difficultto detect by EIA. The use of cell types known to have enhancedbeta-secretase activity, enhanced processing of APP to A beta, and/orenhanced production of A beta are therefore preferred. For example,transfection of cells with the Swedish Mutant form of APP (APP-SW); withAPP-KK; or with APP-SW-KK provides cells having enhanced beta-secretaseactivity and producing amounts of A beta that can be readily measured.

In such assays, for example, the cells expressing APP and beta-secretaseare incubated in a culture medium under conditions suitable forbeta-secretase enzymatic activity at its cleavage site on the APPsubstrate. On exposure of the cells to the compound inhibitor, theamount of A beta released into the medium and/or the amount of CTF99fragments of APP in the cell lysates is reduced as compared with thecontrol. The cleavage products of APP can be analyzed, for example, byimmune reactions with specific antibodies, as discussed above.

Preferred cells for analysis of beta-secretase activity include primaryhuman neuronal cells, primary transgenic animal neuronal cells where thetransgene is APP, and other cells such as those of a stable 293 cellline expressing APP, for example, APP-SW.

In Vivo Assays: Animal Models

Various animal models can be used to analyze beta-secretase activityand/or processing of APP to release A beta, as described above. Forexample, transgenic animals expressing APP substrate and beta-secretaseenzyme can be used to demonstrate inhibitory activity of the compoundsof the invention. Certain transgenic animal models have been described,for example, in U.S. Pat. Nos.: 5,877,399; 5,612,486; 5,387,742;5,720,936; 5,850,003; 5,877,015, and 5,811,633, and in Ganes et.al.,1995, Nature 373:523. Preferred are animals that exhibit characteristicsassociated with the pathophysiology of AD. Administration of thecompound inhibitors of the invention to the transgenic mice describedherein provides an alternative method for demonstrating the inhibitoryactivity of the compounds. Administration of the compounds in apharmaceutically effective carrier and via an administrative route thatreaches the target tissue in an appropriate therapeutic amount is alsopreferred.

Inhibition of beta-secretase mediated cleavage of APP at thebeta-secretase cleavage site and of A beta release can be analyzed inthese animals by measure of cleavage fragments in the animal's bodyfluids such as cerebral fluid or tissues. Analysis of brain tissues forA beta deposits or plaques is preferred.

On contacting an APP substrate with a beta-secretase enzyme in thepresence of an inhibitory compound of the invention and under conditionssufficient to permit enzymatic mediated cleavage of APP and/or releaseof A beta from the substrate, the compounds of the invention areeffective to reduce beta-secretase-mediated cleavage of APP at thebeta-secretase cleavage site and/or effective to reduce released amountsof A beta. Where such contacting is the administration of the inhibitorycompounds of the invention to an animal model, for example, as describedabove, the compounds are effective to reduce A beta deposition in braintissues of the animal, and to reduce the number and/or size of betaamyloid plaques. Where such administration is to a human subject, thecompounds are effective to inhibit or slow the progression of diseasecharacterized by enhanced amounts of A beta, to slow the progression ofAD in the, and/or to prevent onset or development of AD in a patient atrisk for the disease.

Definitions/Abbreviations

The following abbreviations/definitions are used interchangeably herein:

All temperatures are in degrees Celsius (° C.).

TLC refers to thin-layer chromatography.

psi refers to pounds/in².

HPLC refers to high pressure liquid chromatography.

THF refers to tetrahydrofuran.

DMF refers to dimethylformamide.

EDC refers to ethyl-1-(3-dimethylaminopropyl)carbodiimide or1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride.

HOBt refers to 1-hydroxy benzotriazole hydrate.

NMM refers to N-methylmorpholine.

NBS refers to N-bromosuccinimide.

TEA refers to triethylamine.

BOC refers to 1,1-dimethylethoxy carbonyl or t-butoxycarbonyl,—CO—O—C(CH₃)₃.

CBZ refers to benzyloxycarbonyl, —CO—O—CH₂-phenyl.

FMOC refers to 9-fluorenylmethyl carbonate.

TFA refers to trifluoracetic acid, CF₃—COOH.

CDI refers to 1,1′-carbonyldiimidazole.

Saline refers to an aqueous saturated sodium chloride solution.

Chromatography (column and flash chromatography) refers topurification/separation of compounds expressed as (support, eluent). Itis understood that the appropriate fractions are pooled and concentratedto give the desired compound(s).

CMR refers to C-13 magnetic resonance spectroscopy, chemical shifts arereported in ppm (δ) downfield from TMS.

NMR refers to nuclear (proton) magnetic resonance spectroscopy, chemicalshifts are reported in ppm (d) downfield from TMS.

IR refers to infrared spectroscopy.

-phenyl refers to phenyl (C₆H₅).

MS refers to mass spectrometry expressed as m/e, m/z or mass/chargeunit. MH⁺ refers to the positive ion of a parent plus a hydrogen atom.EI refers to electron impact. CI refers to chemical ionization. FABrefers to fast atom bombardment.

HRMS refers to high resolution mass spectrometry.

Ether refers to diethyl ether.

Pharmaceutically acceptable refers to those properties and/or substanceswhich are acceptable to the patient from a pharmacological/toxicologicalpoint of view and to the manufacturing pharmaceutical chemist from aphysical/chemical point of view regarding composition, formulation,stability, patient acceptance and bioavailability.

When solvent pairs are used, the ratios of solvents used arevolume/volume (v/v).

When the solubility of a solid in a solvent is used the ratio of thesolid to the solvent is weight/volume (wt/v).

BOP refers to benzotriazol-1-yloxy-tris(dimethylamino)phosphoniumhexafluorophosphate.

TBDMSCl refers to t-butyldimethylsilyl chloride.

TBDMSOTf refers to t-butyldimethylsilyl trifluosulfonic acid ester.

Trisomy 21 refers to Down's Syndrome.

APP, amyloid precursor protein, is defined as any APP polypeptide,including APP variants, mutations, and isoforms, for example, asdisclosed in U.S. Pat. No. 5,766,846.

A beta, amyloid beta peptide, is defined as any peptide resulting frombeta-secretase mediated cleavage of APP, including peptides of 39, 40,41, 42, and 43 amino acids, and extending from the beta-secretasecleavage site to amino acids 39, 40, 41, 42, or 43.

Beta-secretase (BACE1, Asp2, Memapsin 2) is an aspartyl protease thatmediates cleavage of APP at the amino-terminal edge of A beta. Humanbeta-secretase is described, for example, in WO00/17369.

“Pharmaceutically acceptable” refers to those properties and/orsubstances that are acceptable to the patient from apharmacological/toxicological point of view and to the manufacturingpharmaceutical chemist from a physical/chemical point of view regardingcomposition, formulation, stability, patient acceptance andbioavailability.

A therapeutically effective amount is defined as an amount effective toreduce or lessen at least one symptom of the disease being treated or toreduce or delay onset of one or more clinical markers or symptoms of thedisease.

The present invention provides compounds, compositions, and methods forinhibiting beta-secretase enzyme activity and A beta peptide production.Inhibition of beta-secretase enzyme activity halts or reduces theproduction of A beta from APP and reduces or eliminates the formation ofbeta-amyloid deposits in the brain.

Unless defined otherwise, all scientific and technical terms used hereinhave the same meaning as commonly understood by one of skill in the artto which this invention belongs. The disclosures in this application ofall articles and references, including patents, are incorporated hereinby reference.

The invention is illustrated further by the following examples which arenot to be construed as limiting the invention in scope or spirit to thespecific procedures described in them.

The starting materials and various intermediates may be obtained fromcommercial sources, prepared from commercially available organiccompounds, or prepared using well known synthetic methods.

EXAMPLES

Synthesis

Example A12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione(1)

Step 1: Preparation of{[1-(3-Benzyloxy-5-fluoro-benzyl)-2,3-dihydroxy-propylcarbamoyl]-methyl}-carbamicacid benzyl ester

To a solution of 3-amino-4-(3-benzyloxy-5-fluoro-phenyl)-butane-1,2-diol(5.60 g, 18.34 mmol, 1.00 eq.), triethylamine (5.11 mL, 36.68 mmol, 2.00eq.) and anhydrous DMF (100 mL) at 0° C. is added Z-glycineN-succinimidyl ester (6.18 g, 20.17 mmol, 1.10 eq.) with stirring underN₂. After 2 hours, the reaction is quenched with 1N HCl, extracted withethyl acetate, and washed with 10% NaHCO₃ and then brine. The organiclayer is then dried with MgSO₄, filtered through Celite, andconcentrated in vacuo, yielding the crude product as a light amber oil.Purification via flash chromatography in 5% MeOH/CHCl₃ (rf. 0.33, KMnO₄stain), affords the final product as a white solid (5.40 g., 59% yieldoverall). Calculate mass for C₂₇H₂₉FN₂O₆: 496.20. Mass found forC₂₇H₂₉FN₂O₆: (OAMS) ES+: 497.5 (M+1).

Step 2: Preparation of{[2-(3-Benzyloxy-5-fluoro-phenyl)-1-oxiranyl-ethylcarbamoyl]-methyl}-carbamicacid benzyl ester

In flame-dried glassware is prepared a solution of triphenylphosphine(0.291 g, 1.11 mmol, 1.10 eq.), diisopropylazodicarboxylate (0.22 mL,1.11 mmol, 1.10 eq.) in anh. chloroform (2.5 mL). The product of Step 1is added (0.500 g, 1.01 mmol, 1.00 eq.) with stirring under N₂overnight. The reaction is (complete as monitored by TLC; 35%EtOAc/Hexanes, KMnO₄ stain) concentrated in vacuo to yield crudeproducts as an amber oil. Calculate mass for C₂₇H₂₇FN₂O₅: 478.19. Massfound for C₂₇H₂₇FN₂O₅: (LCMS) ES+: 478.8 (M+1).Step 3: Preparation of{[1-(3-Benzyloxy-5-fluoro-benzyl)-3-(3-ethyl-benzylamino)-2-hydroxy-propylcarbamoyl]-methyl}-carbamicacid benzyl ester

A solution of the product from Step 2 (0.483 g, 1.01 mmol, 1.00 eq.),m-ethyl benzylamine (0.273 g, 2.02 mmol, 2.00 eq.), and isopropanol (5mL) is prepared and heated to 80° C. for 2 hours. The reaction mixtureis then concentrated in vacuo. The resulting crude product is dissolvedin methanol (10 mL) and stirred with Dowex 50WX2-400 ion-exchange resinat 60° C. for 2 hours. The product is released from resin by filteringthrough a frit with 7N NH₃/MeOH. The filtrate is concentrated in vacuoto yield a crude orange product (305 mg, 49% crude yield). The crudeproduct is dissolved in ethyl acetate and washed with 1N HCl, dried withMgSO₄, filtered and concentrated in vacuo. Calculate mass forC₃₆H₄₀FN₃O₅: 613.30. Mass found for C₃₆H-₄₀FN₃O₅: (OAMS) ES+: 614.0(M+1).Step 4: Preparation of[3-(2-Benzyloxycarbonylamino-acetylamino)-4-(3-benzyloxy-5-fluoro-phenyl)-2-hydroxy-butyl]-(3-ethyl-benzyl)-carbamicacid tert-butyl ester

A solution of the product from Step 3 (0.305 g, 0.50 mmol, 1.00 eq.),boc anhydride (0.142 g, 0.65 mmol, 1.30 eq.) and methylene chloride isprepared and stirred under N₂ overnight. The reaction mixture is dilutedwith ethyl acetate, washed with H₂O, and the organic layer issubsequently dried over MgSO₄. Concentration in vacuo affords the crudeproduct as an amber oil (340 mg). The crude product is purified viaflash chromatography using 50% EtOAc/Hexanes as the eluant. The productis concentrated in vacuo yielding a viscous amber oil (133 mg, 0.186mmol; 18% overall yield from the product of Step 1). Calculate mass forC₄₁H₄₈FN₃O₇: 713.35. Mass found for C₄₁H₄₈FN₃O₇: (OAMS) ES+: 713.9(M+1), ES-712.0 (M−1).Step 5: Preparation of[3-(2-Amino-acetylamino)-4-(3-fluoro-5-hydroxy-phenyl)-2-hydroxy-butyl]-(3-ethyl-benzyl)-carbamicacid tert-butyl ester

To a solution of the product from Step 4 (130 mg, 0.18 mmol, 1.00 eq.)in methanol (0.5 mL) is added 10% Pd/C (20 mg, 0.02 mmol, 0.10 eq.). Thereaction vessel is purged with H₂ and then maintained under H₂ via aballoon. The reaction is stirred under H₂ overnight and then filteredand concentrated in vacuo yielding the product as a white solid (63 mg,71% crude yield). Calculate mass for C₂₆H₃₆FN₃O₅: 489.26. Mass found forC₂₆H₃₆FN₃O₅: (OAMS) ES+: 489.8 (M+1), ES-487.8 (M−1).Step 6: Preparation of[3-[2-(5-Bromo-pentanoylamino)-acetylamino]-4-(3-fluoro-5-hydroxy-phenyl)-2-hydroxy-butyl]-(3-ethyl-benzyl)-carbamicacid tert-butyl ester

To a solution of the product from Step 5 (60 mg, 0.12 mmol, 1.00 eq.),triethylamine (33 μL, 0.24 mmol, 2.00 eq.), and anhydrous THF (0.6 mL)is added bromovaleryl chloride (15 μL, 0.11 mmol, 0.95 eq.) under N₂with stirring. The reaction mixture is stirred for 2 hours, then dilutedwith ethyl acetate, quenched with 1N HCl, and washed with 10% NaHCO₃.The organic layer is dried over MgSO₄, filtered, and then concentratedin vacuo, yielding product as a white solid (63 mg, 80% crude yield).Calculate mass for C₃H₄₃BrFN₃O₆: 651.23 (mass calculated for Br⁷⁹isotope). Mass found for C₃₁H₄₃BrFN₃O₆: (LCMS) ES+: 676.1 (M+Na), ES−652.0 (M−1).Step 7: Preparation of12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

To a solution of the product from Step 6 (78 mg, 0.12 mmol, 1.00 eq.)and anhydrous DMF (1 mL) is added Cs₂CO₃ (flame-dried, 78 mg, 0.24 mmol,2.00 eq.) under N₂ with stirring overnight. The reaction mixture isfiltered through Celite and then concentrated in vacuo. The crudeproduct is then stirred with Dowex 50WX2-400 ion-exchange resin at 60°C. for 2 hours. The product is released from resin with 7N NH₃/MeOHthrough a frit and the filtrate is concentrated in vacuo, yielding thefinal product as a white solid (14 mg, 25% yield). Calculate accuratemass for C₂₆H₃₄BrFN₃O₄+H₁: 472.2611. Accurate Mass found forC₂₆H₃₄BrFN₃O₄+H₁: 472.2592.

Compound 1 above is also depicted in Table 1. Compounds 2-125 shownbelow in Table 1 are prepared essentially according to the procedureoutlined in CHARTS A-D and set forth in Example A.

TABLE 1

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

2-{12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl}-acetamide

2-(12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-(12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien- 9-yl)-acetamide

2-{16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl}-acetamide

2-(12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-{12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl}-acetamide

2-(12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-(12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-{16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9- yl}-acetamide

2-(12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-{12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl}-acetamide

2-(12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-(12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-{16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl}-acetamide

2-(12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-{12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl}-acetamide

2-(12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamimo]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-(12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-{16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl}-acetamide

2-(12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide

2-{13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl}-acetamide

2-(13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien- 10-yl)-acetamide

2-(13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien- 10-yl)-acetamide

2-{17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl}-acetamide

2-(13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide

2-{13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl}-acetamide

2-(13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide

2-(13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide

2-{17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl}-acetamide

2-(13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene- 7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15- triene-7,10-dione

13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene- 8,11-dione

13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16- triene-8,11-dione

17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene- 8,11-dione

13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16- triene-8,11-dione

13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene- 8,11-dione

13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene- 7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene- 7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene- 7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene- 7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15- triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene- 7,10-dione

12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15- triene-7,10-dione

12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15- triene-7,10-dione

12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione

13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene- 8,11-dione

13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene- 8,11-dione

13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16- triene-8,11-dione

13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene- 8,11-dione

13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione

BIOLOGICAL EXAMPLES Example A

Enzyme Inhibition Assay

The compounds of the invention are analyzed for inhibitory activity byuse of the MBP-C125 assay. This assay determines the relative inhibitionof beta-secretase cleavage of a model APP substrate, MBP-C125SW, by thecompounds assayed as compared with an untreated control. A detaileddescription of the assay parameters can be found, for example, in U.S.Pat. No. 5,942,400. Briefly, the substrate is a fusion peptide formed ofmaltose binding protein (MBP) and the carboxy terminal 125 amino acidsof APP-SW, the Swedish mutation. The beta-secretase enzyme is derivedfrom human brain tissue as described in Sinha et.al, 1999, Nature40:537-540) or recombinantly produced as the full-length enzyme (aminoacids 1-501), and can be prepared, for example, from 293 cellsexpressing the recombinant cDNA, as described in WO00/47618.

Inhibition of the enzyme is analyzed, for example, by immunoassay of theenzyme's cleavage products. One exemplary ELISA uses an anti-MBP captureantibody that is deposited on precoated and blocked 96-well high bindingplates, followed by incubation with diluted enzyme reaction supernatant,incubation with a specific reporter antibody, for example, biotinylatedanti-SW192 reporter antibody, and further incubation withstreptavidin/alkaline phosphatase. In the assay, cleavage of the intactMBP-C125SW fusion protein results in the generation of a truncatedamino-terminal fragment, exposing a new SW-192 antibody-positive epitopeat the carboxy terminus. Detection is effected by a fluorescentsubstrate signal on cleavage by the phosphatase. ELISA only detectscleavage following Leu 596 at the substrate's APP-SW 751 mutation site.

Specific Assay Procedure

Compounds are diluted in a 1:1 dilution series to a six-pointconcentration curve (two wells per concentration) in one 96-plate rowper compound tested. Each of the test compounds is prepared in DMSO tomake up a 10 millimolar stock solution. The stock solution is seriallydiluted in DMSO to obtain a final compound concentration of 200micromolar at the high point of a 6-point dilution curve. Ten (10)microliters of each dilution is added to each of two wells on row C of acorresponding V-bottom plate to which 190 microliters of 52 millimolarNaOAc, 7.9% DMSO, pH 4.5 are pre-added. The NaOAc diluted compound plateis spun down to pellet precipitant and 20 microliters/well istransferred to a corresponding flat-bottom plate to which 30 microlitersof ice-cold enzyme-substrate mixture (2.5 microliters MBP-C125SWsubstrate, 0.03 microliters enzyme and 24.5 microliters ice cold 0.09%TX100 per 30 microliters) is added. The final reaction mixture of 200micromolar compound at the highest curve point is in 5% DMSO, 20millimolar NaAc, 0.06% TX100, at pH 4.5.

Warming the plates to 37 degrees C. starts the enzyme reaction. After 90minutes at 37 degrees C., 200 microliters/well cold specimen diluent isadded to stop the reaction and 20 microliters/well is transferred to acorresponding anti-MBP antibody coated ELISA plate for capture,containing 80 microliters/well specimen diluent. This reaction isincubated overnight at 4 degrees C. and the ELISA is developed the nextday after a 2 hours incubation with anti-192SW antibody, followed byStreptavidin-AP conjugate and fluorescent substrate. The signal is readon a fluorescent plate reader.

Relative compound inhibition potency is determined by calculating theconcentration of compound that showed a fifty percent reduction indetected signal (IC₅₀) compared to the enzyme reaction signal in thecontrol wells with no added compound. In this assay, the compounds ofthe invention exhibited an IC₅₀ of less than 50 micromolar.

Example B

Cell Free Inhibition Assay Utilizing a Synthetic APP Substrate

A synthetic APP substrate that can be cleaved by beta-secretase andhaving N-terminal biotin and made fluorescent by the covalent attachmentof Oregon green at the Cys residue is used to assay beta-secretaseactivity in the presence or absence of the inhibitory compounds of theinvention. Substrates include the following:

-   Biotin-SEVNL-DAEFRC[oregon green]KK [SEQ ID NO: 1]-   Biotin-SEVKM-DAEFRC[oregon green]KK [SEQ ID NO: 2]-   Biotin-GLNIKTEEISEISY-EVEFRC[oregon green]KK [SEQ ID NO: 3]-   Biotin-ADRGLTTRPGSGLTNIKTEEISEVNL-DAEFRC[oregon green]KK [SEQ ID    NO:4]-   Biotin-FVNQHLCoxGSHLVEALY-LVCoxGERGFFYTPKAC[oregon green]KK [SEQ ID    NO: 5]

The enzyme (0.1 nanomolar) and test compounds (0.001-100 micromolar) areincubated in pre-blocked, low affinity, black plates (384 well) at 37degrees C. for 30 minutes. The reaction is initiated by addition of 150millimolar substrate to a final volume of 30 microliter per well. Thefinal assay conditions are: 0.001-100 micromolar compound inhibitor; 0.1molar sodium acetate (pH 4.5); 150 nanomolar substrate; 0.1 nanomolarsoluble beta-secretase; 0.001% Tween 20, and 2% DMSO. The assay mixtureis incubated for 3 hours at 37 degrees C., and the reaction isterminated by the addition of a saturating concentration of immunopurestreptavidin. After incubation with streptavidin at room temperature for15 minutes, fluorescence polarization is measured, for example, using aLJL Acqurest (Ex485 nm/Em530 nm). The activity of the beta-secretaseenzyme is detected by changes in the fluorescence polarization thatoccur when the substrate is cleaved by the enzyme. Incubation in thepresence or absence of compound inhibitor demonstrates specificinhibition of beta-secretase enzymatic cleavage of its synthetic APPsubstrate. In this assay, compounds of the invention exhibited an IC50of less than 50 micromolar.

Example C

Beta-Secretase Inhibition: P26-P4′ SW Assay

Synthetic substrates containing the beta-secretase cleavage site of APPare used to assay beta-secretase activity, using the methods described,for example, in published PCT application WO00/47618. The P26-P4′ SWsubstrate is a peptide of the sequence:(biotin)CGGADRGLTTRPGSGLTNIKTEEISEVNLDAEF [SEQ ID NO: 6] The P26-P1standard has the sequence: (biotin)CGGADRGLTTRPGSGLTNIKTEEISEVNL [SEQ IDNO: 7]

Briefly, the biotin-coupled synthetic substrates are incubated at aconcentration of from about 0 to about 200 micromolar in this assay.When testing inhibitory compounds, a substrate concentration of about1.0 micromolar is preferred. Test compounds diluted in DMSO are added tothe reaction mixture, with a final DMSO concentration of 5%. Controlsalso contain a final DMSO concentration of 5%. The concentration of betasecretase enzyme in the reaction is varied, to give productconcentrations with the linear range of the ELISA assay, about 125 to2000 picomolar, after dilution.

The reaction mixture also includes 20 millimolar sodium acetate, pH 4.5,0.06% Triton X100, and is incubated at 37 degrees C. for about 1 to 3hours. Samples are then diluted in assay buffer (for example, 145.4nanomolar sodium chloride, 9.51 millimolar sodium phosphate, 7.7millimolar sodium azide, 0.05% Triton X405, 6 g/liter bovine serumalbumin, pH 7.4) to quench the reaction, then diluted further forimmunoassay of the cleavage products.

Cleavage products can be assayed by ELISA. Diluted samples and standardsare incubated in assay plates coated with capture antibody, for example,SW192, for about 24 hours at 4 degrees C. After washing in TTBS buffer(150 millimolar sodium chloride, 25 millimolar Tris, 0.05% Tween 20, pH7.5), the samples are incubated with strepavidin-AP according to themanufacturer's instructions. After a one hour incubation at roomtemperature, the samples are washed in TTBS and incubated withfluorescent substrate solution A (31.2 g/liter2-amino-2-methyl-1-propanol, 30 mg/liter, pH 9.5). Reaction withstreptavidin-alkaline phosphate permits detection by fluorescence.Compounds that are effective inhibitors of beta-secretase activitydemonstrate reduced cleavage of the substrate as compared to a control.

Example D

Assays using Synthetic Oligopeptide-Substrates

Synthetic oligopeptides are prepared that incorporate the known cleavagesite of beta-secretase, and optionally detectable tags, such asfluorescent or chouromogenic moieties. Examples of such peptides, aswell as their production and detection methods are described in U.S.Pat. No. 5,942,400, herein incorporated by reference. Cleavage productscan be detected using high performance liquid chromatography, orfluorescent or chromogenic detection methods appropriate to the peptideto be detected, according to methods well known in the art.

By way of example, one such peptide has the sequence SEVNL-DAEF [SEQ IDNO: 8], and the cleavage site is between residues 5 and 6. Anotherpreferred substrate has the sequence ADRGLTTRPGSGLTNIKTEEISEVNL-DAEF[SEQ ID NO: 9], and the cleavage site is between residues 26 and 27.

These synthetic APP substrates are incubated in the presence ofbeta-secretase under conditions sufficient to result in beta-secretasemediated cleavage of the substrate. Comparison of the cleavage resultsin the presence of the compound inhibitor to control results provides ameasure of the compound's inhibitory activity.

Example E

Inhibition of Beta-Secretase Activity—Cellular Assay

An exemplary assay for the analysis of inhibition of beta-secretaseactivity utilizes the human embryonic kidney cell line HEKp293 (ATCCAccession No. CRL-1573) transfected with APP751 containing the naturallyoccurring double mutation Lys651Met52 to Asn651Leu652 (numbered forAPP751), commonly called the Swedish mutation and shown to overproduce Abeta (Citron et.al., 1992, Nature 360:672-674), as described in U.S.Pat. No. 5,604,102.

The cells are incubated in the presence/absence of the inhibitorycompound (diluted in DMSO) at the desired concentration, generally up to10 micrograms/ml. At the end of the treatment period, conditioned mediais analyzed for beta-secretase activity, for example, by analysis ofcleavage fragments. A beta can be analyzed by immunoassay, usingspecific detection antibodies. The enzymatic activity is measured in thepresence and absence of the compound inhibitors to demonstrate specificinhibition of beta-secretase mediated cleavage of APP substrate.

Example F

Inhibition of Beta-Secretase in Animal Models of AD

Various animal models can be used to screen for inhibition ofbeta-secretase activity. Examples of animal models useful in theinvention include, but are not limited to, mouse, guinea pig, dog, andthe like. The animals used can be wild type, transgenic, or knockoutmodels. In addition, mammalian models can express mutations in APP, suchas APP695-SW and the like described herein. Examples of transgenicnon-human mammalian models are described in U.S. Pat. Nos. 5,604,102,5,912,410 and 5,811,633.

PDAPP mice, prepared as described in Games et.al., 1995, Nature373:523-527 are useful to analyze in vivo suppression of A beta releasein the presence of putative inhibitory compounds. As described in U.S.Pat. No. 6,191,166, 4 month old PDAPP mice are administered compoundformulated in vehicle, such as corn oil. The mice are dosed withcompound (1-30 mg/ml; preferably 1-10 mg/ml). After time, e.g., 3-10hours, the animals are sacrificed, and brains removed for analysis.

Transgenic animals are administered an amount of the compound inhibitorformulated in a carrier suitable for the chosen mode of administration.Control animals are untreated, treated with vehicle, or treated with aninactive compound. Administration can be acute, i.e., single dose ormultiple doses in one day, or can be chronic, i.e., dosing is repeateddaily for a period of days. Beginning at time 0, brain tissue orcerebral fluid is obtained from selected animals and analyzed for thepresence of APP cleavage peptides, including A beta, for example, byimmunoassay using specific antibodies for A beta detection. At the endof the test period, animals are sacrificed and brain tissue or cerebralfluid is analyzed for the presence of A beta and/or beta-amyloidplaques. The tissue is also analyzed for necrosis.

Animals administered the compound inhibitors of the invention areexpected to demonstrate reduced A beta in brain tissues or cerebralfluids and reduced beta amyloid plaques in brain tissue, as comparedwith non-treated controls.

Example G

Inhibition of A Beta Production in Human Patients

Patients suffering from Alzheimer's Disease (AD) demonstrate anincreased amount of A beta in the brain. AD patients are administered anamount of the compound inhibitor formulated in a carrier suitable forthe chosen mode of administration. Administration is repeated daily forthe duration of the test period. Beginning on day 0, cognitive andmemory tests are performed, for example, once per month.

Patients administered the compound inhibitors are expected todemonstrate slowing or stabilization of disease progression as analyzedby changes in one or more of the following disease parameters: A betapresent in CSF or plasma; brain or hippocampal volume; A beta depositsin the brain; amyloid plaque in the brain; and scores for cognitive andmemory function, as compared with control, non-treated patients.

Example H

Prevention of A Beta Production in Patients at Risk for AD

Patients predisposed or at risk for developing AD are identified eitherby recognition of a familial inheritance pattern, for example, presenceof the Swedish Mutation, and/or by monitoring diagnostic parameters.Patients identified as predisposed or at risk for developing AD areadministered an amount of the compound inhibitor formulated in a carriersuitable for the chosen mode of administration. Administration isrepeated daily for the duration of the test period. Beginning on day 0,cognitive and memory tests are performed, for example, once per month.

Patients administered the compound inhibitors are expected todemonstrate slowing or stabilization of disease progression as analyzedby changes in one or more of the following disease parameters: A betapresent in CSF or plasma; brain or hippocampal volume; amyloid plaque inthe brain; and scores for cognitive and memory function, as comparedwith control, non-treated patients.

The invention and the manner and process of making and using it, are nowdescribed in such full, clear, concise and exact terms as to enable anyperson skilled in the art to which it pertains, to make and use thesame. It is to be understood that the foregoing describes preferredembodiments of the present invention and that modifications may be madetherein without departing from the spirit or scope of the presentinvention as set forth in the claims. To particularly point out anddistinctly claim the subject matter regarded as invention, the followingclaims conclude this specification.

1. A compound of the formula:

or a pharmaceutically acceptable salt thereof, wherein U is

- - - is an optional bond; J is —CH₂OH or —NH—R_(c) when - - - is not abond, or absent when - - - is a bond; G is OH when - - - is not a bondor —O— when - - - is a bond; R is hydrogen or C₁-C₆ alkyl; m is 1-6; R₄and R₅ are independently H, C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl,C₁-C₆ haloalkyl, C₃-C₇ cycloalkyl, C₄-C₁₂ cycloalkylalkyl, C₁-C₆alkoxyalkyl, or C₃-C₆ cycloalkyl; X represents —(CR₄R₅)_(m)—; Y ishydrogen, C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, C₁-C₆ haloalkyl,C₃-C₇ cycloalkyl, C₄-C₁₂ cycloalkylalkyl, C₁-C₆ alkoxyalkyl, C₃-C₆cycloalkyl, or Y together with the carbon to which it is attached is a Dor L amino acid side chain; R₆, R_(6′) and R_(6″) independently areC₁-C₆ alkyl optionally substituted with one, two or three groupsindependently selected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃,C₁-C₃ alkoxy, amino, and mono- or dialkylamino; or C₂-C₆ alkenyl orC₂-C₆ alkynyl, each of which is optionally substituted with one, two orthree groups independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,—C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino; or—(CH₂)₀₋₄—O—(C₁-C₆ alkyl), where the alkyl portion is optionallysubstituted with one, two, three, four, or five groups independentlyselected from halogen; or —OH, —NO₂, halogen, —CO₂H, —C≡N,—(CH₂)₀₋₄—CO—NR₈R₉, —(CH₂)₀₋₄—CO—(C₁-C₁₂ alkyl), —(CH₂)₀₋₄—CO—(C₂-C₁₂alkenyl), —(CH₂)₀₋₄—CO—(C₂-C₁₂ alkynyl), —(CH₂)₀₋₄—CO—(C₃-C-7cycloalkyl), —(CH₂)₀₋₄—R_(aryl), —CH₂)₀₋₄R_(heteroaryl),—(CH₂)₀₋₄—R_(heterocyclyl), —(CH₂)₀₋₄—CO—R_(aryl),—(CH₂)₀₋₄—CO—R_(heteroaryl), —(CH₂)₀₋₄—CO—R_(heterocyclyl),—(CH₂)₀₋₄—CO—R₁₀, —(CH₂)₀₋₄—CO—O—R₁₁, —(CH₂)₀₋₄—SO₂—NR₈R₉,—(CH₂)₀₋₄—SO—(C₁-C₈ alkyl), —(CH₂)₀₋₄—SO₂—(C₁-C₁₂ alkyl),—(CH₂)₀₋₄—SO₂—(C₃-C₇ cycloalkyl), —(CH₂)₀₋₄—N(H or R₁₁)—CO—O—R₁₁,—(CH₂)₀₋₄—N(H or R₁₁)—CO—N(R₁₁)₂, —(CH₂)₀₋₄—N—CS—N(R₁₁)₂, —(CH₂)₀₋₄—N(—Hor R₁₁)—CO—R₈, —(CH₂)₀₋₄—NR₈R₉, —(CH₂)₀₋₄—R₁₀, —(CH₂)₀₋₄—O—CO—(C₁-C₆alkyl), —(CH₂)₀₋₄—O—P(O)—(O—R_(aryl))₂, —(CH₂)₀₋₄—O—CO—N(R₁₁)₂,—(CH₂)₀₋₄—O—CS—N(R₁₁)₂, —(CH₂)₀₋₄—O—(R₁₁), —(CH₂)₀₋₄—O—(R₁₁)—COOH,—(CH₂)₀₋₄—S—(R₁₁), C₃-C₇ cycloalkyl, —(CH₂)₀₋₄—N(—H or R₁₁)—SO₂—R₇, or—(CH₂)₀₋₄—C₃-C₇ cycloalkyl; R₈ and R₉ are the same or different andrepresent —H, —C₃-C₇ cycloalkyl, —(C₁-C₂ alkyl)-(C₃-C₇ cycloalkyl),—(C₁-C₆ alkyl)—O—(C₁-C₃ alkyl), —C₁-C₆ alkenyl, —C₁-C₆ alkynyl, or—C₁-C₆ alkyl chain with one double bond and one triple bond; or —C₁-C₆alkyl optionally substituted with —OH or —NH₂; or —C₁-C₆ alkyloptionally substituted with one, two or three groups independentlyselected from halogen; or heterocyclyl optionally substituted with one,two or three groups selected from halogen, amino, mono- or dialkylamino,—OH, —C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆ alkyl, —SO₂—N(C₁-C₆ alkyl)₂,—SO₂—(C₁-C₄ alkyl), —CO—NH₂, —CO—NH—C₁-C₆ alkyl, oxo, —CO—N(C₁-C₆alkyl)₂, C₁-C₆ alkyl optionally substituted with one, two or threegroups independently selected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N,—CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino, C₂-C₆ alkenyl orC₂-C₆ alkynyl, each of which is optionally substituted with one, two orthree groups independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,—C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylamino, and C₁-C₆alkoxy optionally substituted with one, two or three groupsindependently selected from halogen; or aryl or heteroaryl, each ofwhich is optionally substituted with one, two or three groupsindependently selected from halogen, amino, mono- or dialkylamino, —OH,—C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆ alkyl, —SO₂—N(C₁-C₆ alkyl)₂, —SO₂—(C₁-C₄alkyl), —O—NH₂, —CO—NH—C₁-C₆ alkyl, and —CO—N(C₁-C₆ alkyl)₂, C₁-C₆ alkyloptionally substituted with one, two or three groups independentlyselected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,amino, and mono- or dialkylamino, C₂-C₆ alkenyl or C₂-C₆ alkynyl, eachof which is optionally substituted with one, two or three groupsindependently selected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃,C₁-C₃ alkoxy, amino, and mono- or dialkylamino, and C₁-C₆ alkoxyoptionally substituted with one, two or three of halogen; R₁₀ isheterocyclyl optionally substituted with one, two, three or four groupsindependently selected from C₁-C₆ alkyl; R₁₁ is C₁-C₆ alkyl, C₂-C₆alkenyl, C₂-C₆ alkynyl, C₃-C₇ cycloalkyl, —(CH₂)₀₋₂—R_(aryl), or—(CH₂)₀₋₂—R_(heteroaryl); R_(aryl) is aryl optionally substituted withone, two or three groups independently selected from halogen, amino,mono- or dialkylamino, —OH, —C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆ alkyl,—SO₂—N(C₁-C₆ alkyl)₂, —SO₂—(C₁-C₄ alkyl), —CO—NH₂, —CO—NH—C₁-C₆ alkyl,—CO—N(C₁-C₆ alkyl)₂, C₁-C₆ alkyl optionally substituted with one, two orthree groups independently selected from C₁-C₃ alkyl, halogen, —OH, —SH,—C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- or dialkylaniino, C₂-C₆alkenyl or C₂-C₆ alkynyl, each of which is optionally substituted withone, two or three groups independently selected from C₁-C₃ alkyl,halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy, amino, and mono- ordialkylamino, and C₁-C₆ alkoxy optionally substituted with one, two orthree groups independently selected from halogen; R_(heteroaryl) isheteroaryl optionally substituted with one, two or three groupsindependently selected from halogen, amino, mono- or dialkylaniino, —OH,—C≡N, —SO₂—NH₂, —SO₂—NH—C₁-C₆ alkyl, —SO₂—N(C₁-C₆ alkyl)₂, —SO₂—(C₁-C₄alkyl), —CO—N₂, —CO—NH—C₁-C₆ alkyl, —CO—N(C₁-C₆ alkyl)₂, C₁-C₆ alkyloptionally substituted with one, two or three groups independentlyselected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,amino, and mono- or dialkylamino, C₂-C₆ alkenyl or C₂-C₆ alkynyl, eachof which is optionally substituted with one, two or three groupsindependently selected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃,C₁-C₃ alkoxy, amino, and mono- or dialkylamino, and C₁-C₆ alkoxyoptionally substituted with one, two or three groups independentlyselected from halogen; R_(heterocyclyl) is heterocyclyl optionallysubstituted with one, two or three groups independently selected fromhalogen, amino, mono- or dialkylamino, —OH, —C≡N, —SO₂—NH₂,—SO₂—NH—C₁-C₆ alkyl, —SO₂—N(C₁-C₆ alkyl)₂, —SO₂—(C₁-C₄ alkyl), —CO—NH₂,—CO—NH—C₁-C₆ alkyl, ═O, —CO—N(C₁-C₆ alkyl)₂, C₁-C₆ alkyl optionallysubstituted with one, two or three groups independently selected fromC₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy, amino, andmono- or dialkylamino, C₂-C₆ alkenyl or C₂-C₆ alkynyl, each of which isoptionally substituted with one, two or three groups independentlyselected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,amino, and mono- or dialkylamino, and C₁-C₆ alkoxy optionallysubstituted with one, two or three groups independently selected fromhalogen; R₂ is —H; or —(CH₂)₀₋₄—R_(aryl) and —(CH₂)₀₋₄—R_(heteroaryl);or C₁-C₆ alkyl optionally substituted with one, two or three groupsindependently selected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃,C₁-C₃ alkoxy, amino, and mono- or dialkylamino; or C₂-C₆ alkenyl, C₂-C₆alkynyl or —(CH₂)₀₋₄—C₃-C₇ cycloalkyl, each of which is optionallysubstituted with one, two or three groups independently selected fromC₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy, amino, andmono- or dialkylamino; R₃ is —H, C₂-C₆ alkenyl, C₂-C₆ alkynyl,—(CH₂)₀₋₄—R_(aryl), or (CH₂)₀₋₄—R_(heteroaryl); or C₁-C₆ alkyloptionally substituted with one, two or three groups independentlyselected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,amino, and mono- ordialkylamino; or —(CH₂)₀₋₄—C₃-C₇ cycloalkyloptionally substituted with one, two or three groups independentlyselected from C₁-C₃ alkyl, halogen, —OH, —SH, —C≡N, —CF₃, C₁-C₃ alkoxy,amino, and mono- or dialkylamino; or R₂ and R₃ taken together with thecarbon atom to which they are attached form a carbocycle of three, four,five, six, or seven carbon atoms, where one atom is optionally aheteroatom selected from the group consisting of —O—, —S—, —SO₂—, and—NR₈—; R_(C) hydrogen, —(CR₂₄₅R₂₅₀)₀₋₄-aryl, —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl,—(CR₂₄₅R₂₅₀)₀₋₄-heterocyclyl, —(CR₂₄₅R₂₅₀)₀₋₄-aryl-heteroaryl,—(CR₂₄₅R₂₅₀)₀₋₄-aryl-heterocyclyl, —(CR₂₄₅R₂₅₀)₀₋₄-aryl-aryl,—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl-aryl,—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl-heterocyclyl,—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl-heteroaryl,—(CR₂₄₅R₂₅₀)₀₋₄-heterocyclyl-heteroaryl,—(CR₂₄₅R₂₅₀)₀₋₄-heterocyclyl-heterocyclyl,—(CR₂₄₅R₂₅₀)₀₋₄-heterocyclyl-aryl, —[C(R₂₅₅)(R₂₆₀)]₁₋₃—CO—N—(R₂₅₅)₂,—CH(aryl)₂, —CH(heteroaryl)₂, —CH(heterocyclyl)₂, —CH(aryl)(heteroaryl),—(CH₂)₀₋₁—CH((CH₂)₀₋₆—OH)—(CH₂)₀₋₁-aryl,—(CH₂)₀₋₁—CH((CH₂)₀₋₆—OH—(CH₂)₀₋₁-heteroaryl, —CH(-aryl or-heteroaryl)-CO—O(C₁-C₄ alkyl), —CH(—CH₂—OH)—CH(OH)-phenyl-NO₂, (C₁-C₆alkyl)-O—(C₁-C₆ alkyl)—OH; —CH₂—NH—CH₂-CH(—O—CH₂—CH₃)₂,—(CH₂)₀₋₆—C(═NR₂₃₅)(NR₂₃₅R₂₄₀), or —(CH₂)₀₋₃—(C₃-C₈) cycloalkyl whereinthe cycloalkyl is optionally substituted with 1, 2, or 3 groupsindependently selected from the group consisting of R₂₀₅, —CO₂H, and—CO₂—(C₁-C₄ alkyl), or cyclopentyl, cyclohexyl, or cycloheptyl ringfused to aryl, heteroaryl, or heterocyclyl wherein one, two or threecarbons of the cyclopentyl, cyclohexyl, or cycloheptyl is optionallyreplaced with a heteroatom independently selected from NH, NR₂₁₅, O, orS(═O)₀₋₂, and wherein the cyclopentyl, cyclohexyl, or cycloheptyl groupcan be optionally substituted with one or two groups that areindependently R₂₀₅, ═O, —CO—NR₂₃₅R₂₄₀, or —SO₂—(C₁-C₄ alkyl), or C₂-C₁₀alkenyl or C₂-C₁₀ alkynyl, each of which is optionally substituted with1, 2, or 3 R₂₀₅ groups, wherein each aryl and heteroaryl is optionallysubstituted with 1, 2, or 3 R₂₀₀, and wherein each heterocyclyl isoptionally substituted with 1, 2, 3, or 4 R₂₁₀; R₂₀₀ at each occurrenceis independently selected from —OH, —NO₂, halogen, —CO₂H, C≡N,—(CH₂)₀₋₄—CO—NR₂₂₀R₂₂₅, —(CH₂)₀₋₄—CO—(C₁-C₁₂ alkyl),—(CH₂)₀₋₄—CO—(C₂-C₁₂ alkenyl), —(CH₂)₀₋₄—CO—(C₂-C₁₂ alkynyl),—(CH₂)₀₋₄—CO—(C₃-C₇ cycloalkyl), —(CH₂)₀₋₄—CO-aryl,—(CH₂)₀₋₄—CO-heteroaryl, —(CH₂)₀₋₄—CO-heterocyclyl, —(CH₂)₀₋₄—CO—O—R₂₁₅,—(CH₂)₀₋₄SO₂—NR₂₂₀R₂₂₅, —(CH₂)₀₋₄—SO—(C₁-C₈ alkyl), —(CH₂)₀₋₄—SO₂(C₁-C₁₂alkyl), —(CH₂)₀₋₄—SO₂—(C₃-C₇ cycloalkyl), —(CH₂)₀₋₄—N(H orR₂₁₅)—CO—O—R₂₁₅, —(CH₂)₀₋₄—N(H or R₂₁₅)—CO—N(R₂₁₅)₂,—(CH₂)₀₋₄—N—CS—N(R₂₁₅)₂, —(CH₂)₀₋₄—N(—H or R₂₁₅)—CO—R₂₂₀,—(CH₂)₀₋₄—NR₂₂₀R₂₂₅, —(CH₂)₀₋₄—O—CO—(C₁-C₆ alkyl),—(CH₂)₀₋₄—O—P(O)—(OR₂₄₀)₂, —(CH₂)₀₋₄—O—CO—N(R₂₁₅)₂,—(CH₂)₀₋₄—O—CS—N(R₂₁₅)₂, —(CH₂)₀₋₄—O—(R₂₁₅), —(CH₂)₀₋₄—O—(R₂₁₅)—COOH,—(CH₂)_(0-4 —S—(R) ₂₁₅), —(CH₂)₀₋₄—O—(C₁-C₆ alkyl optionally substitutedwith 1, 2, 3, or 5 —F), C₃-C₇ cycloalkyl, —(CH₂)₀₋₄—N(H orR₂₁₅)—SO₂—R₂₂₀, —(CH₂)₀₋₄—C₃-C₇ cycloalkyl, or C₁-C₁₀ alkyl optionallysubstituted with 1, 2, or 3 R₂₀₅ groups, or C₂-C₁₀ alkenyl or C₂-C₁₀alkynyl, each of which is optionally substituted with 1 or 2 R₂₀₅groups, wherein the aryl and heteroaryl groups at each occurrence areoptionally substituted with 1, 2, or 3 groups that are independentlyR₂₀₅, R₂₁₀, or C₁-C₆ alkyl substituted with 1, 2, or 3 groups that areindependently R₂₀₅ or R₂₁₀, and wherein the heterocyclyl group at eachoccurrence is optionally substituted with 1, 2, or 3 groups that areindependently R₂₁₀; R₂₀₅ at each occurrence is independently selectedfrom C₁-C₆ alkyl, halogen, —OH, —O-phenyl, —SH, —C≡N, —CF₃, C₁-C₆alkoxy, NH₂, NH(C₁-C₆ alkyl) or N—(C₁-C₆ alkyl)(C₁-C₆ alkyl); R₂₁₀ ateach occurrence is independently selected from halogen, C₁-C₆ alkoxy,C₁-C₆ haloalkoxy, —NR₂₂₀R₂₂₅, OH, C≡N, —CO—(C₁-C₄ alkyl),_SO_(2—)NR₂₃₅R₂₄₀, —CO—NR₂₃₅R₂₄₀, —SO₂—(C₁-C₄ alkyl), ═O, or C₁-C₆alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl or C₃-C₇ cycloalkyl, each of whichis optionally substituted with 1, 2, or 3 R₂₀₅ groups; R₂₁₅ at eachoccurrence is independently selected from C₁-C₆ alkyl, —(CH₂)₀₋₂-(aryl),C₂-C₆ alkenyl, C₂-C₆ alkynyl, C₃-C₇ cycloalkyl, and—(CH₂)₀₋₂-(heteroaryl), —(CH₂)₀₋₂-(heterocyclyl), wherein the aryl groupat each occurrence is optionally substituted with 1, 2, or 3 groups thatare independently R₂₀₅ or R₂₁₀, and wherein the heterocyclyl andheteroaryl groups at each occurrence are optionally substituted with 1,2, or 3 R₂₁₀; R₂₂₀ and R₂₂₅ at each occurrence are independentlyselected from —H, —C₃-C₇ cycloalkyl, —(C₁-C₂ alkyl)-(C₃-C₇ cycloalkyl),—(C₁-C₆ alkyl)—O—(C₁-C₃ alkyl), —C₂-C₆ alkenyl, —C₂-C₆ alkynyl —C₁-C₆alkyl chain with one double bond and one triple bond, -aryl,-heteroaryl, and -heterocyclyl, or —C₁-C₁₀ alkyl optionally substitutedwith —OH, —NH₂ or halogen, wherein the aryl, heterocyclyl and heteroarylgroups at each occurrence are optionally substituted with 1, 2, or 3R₂₇₀ groups R₂₃₅ and R₂₄₀ at each occurrence are independently H, orC₁-C₆ alkyl; R₂₄₅ and R₂₅₀ at each occurrence are independently selectedfrom —H, C₁-C₄ alkyl, C₁-C₄ alkylaryl, C₁-C₄ alkylheteroaryl, C₁-C₄hydroxyalkyl, C₁-C₄ alkoxy, C₁-C₄ haloalkoxy, —(CH₂)₀₋₄—C₃-C₇cycloalkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl, and phenyl; or R₂₄₅ and R₂₅₀are taken together with the carbon to which they are attached to form acarbocycle of 3, 4, 5, 6, or 7 carbon atoms, where one carbon atom isoptionally replaced by a heteroatom selected from —O—, —S—, —SO₂—, and—NR₂₂₀—; R₂₅₅ and R₂₆₀ at each occurrence are independently selectedfrom —H, —(CH₂)₁₋₂—S(O)₀₋₂—(C₁-C₆ alkyl), —(C₁-C₄ alkyl)-aryl, —(C₁-C₄alkyl)-heteroaryl, —(C₁-C₄ alkyl)-heterocyclyl, -aryl, -heteroaryl,-heterocyclyl, —(CH₂)₁₋₄—R₂₆₅—(CH₂)₀₋₄-aryl,—(CH₂)₁₋₄—R₂₆₅—(CH₂)₀₋₄-heteroaryl,—(CH₂)₁₋₄—R₂₆₅—(CH₂)₀₋₄-heterocyclyl, or C₁-C₆ alkyl, C₂-C₆ alkenyl,C₂-C₆ alkynyl or —(CH₂)₀₋₄—C₃-C₇ cycloalkyl, each of which is optionallysubstituted with 1, 2, or 3 R₂₀₅ groups, wherein each aryl or phenyl isoptionally substituted with 1, 2, or 3 groups that are independentlyR₂₀₅, R₂₁₀, or C₁-C₆ alkyl substituted with 1, 2, or 3 groups that areindependently R₂₀₅ or R₂₁₀, and wherein each heterocyclyl is optionallysubstituted with 1, 2, 3, or 4 R₂₁₀; R₂₆₅ at each occurrence isindependently —O—, —S— or —N(C₁-C₆ alkyl)-; R₂₇₀ at each occurrence isindependently R₂₀₅, halogen C₁-C₆ alkoxy, C₁-C₆ haloalkoxy, NR₂₃₅R₂₄₀,—OH, —C≡N, —CO—(C₁-C₄ alkyl), _SO_(2—)NR₂₃₅R₂₄₀, —CO—NR₂₃₅R₂₄₀,—SO₂—(C₁-C₄ alkyl), ═O, or C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkynyl or—(CH₂)₀₋₄—C₃-C₇ cycloalkyl, each of which is optionally substituted with1, 2, or 3 R₂₀₅ groups.
 2. A compound according to claim 1 having theformula


3. A compound according to claim 2 wherein Y is alkynyl, or Y togetherwith the carbon to which it is attached is a D or L amino acid sidechain; X is C₁-C₆ alkyl; R₂ and R₃ are hydrogen; and R_(c) is—(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.
 4. A compound according toclaim 3 wherein Y is hydrogen, alkynyl, —CH(CH₃)CH₃ or —CH₂CH₂SCH₃; andR_(c) is phenylmethyl, pyridin-3-ylmethyl, phenylcyclpropyl orpyridin-3-ylcyclpropyl optionally substituted with C₁-C₆ alkyl, C₂-C₆alkynyl or trifluoromethyl.
 5. A compound according to claim 2 havingthe formula


6. A compound according to claim 5 wherein Y is alkynyl, or Y togetherwith the carbon to which it is attached is a D or L amino acid sidechain; X is C₁-C₆ alkyl; R₂ and R₃ are hydrogen; and R_(c) is—(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.
 7. A compound according toclaim 1 having the formula


8. A compound according to claim 7 wherein X is C₁-C₆ alkyl; R₂ and R₃are hydrogen; and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which is optionally substituted withone or two R₂₀₀.
 9. A compound according to claim 1 having the formula:


10. A compound according to claim 9 wherein X is C₁-C₆ alkyl; R₂ and R₃are hydrogen; and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which is optionally substituted withone or two R₂₀₀.
 11. A compound according to claim 1 having the formula:


12. A compound according to claim 11 wherein X is C₁-C₆alkyl; R₂ and R₃are hydrogen; and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which is optionally substituted withone or two R₂₀₀.
 13. A compound according to claim 1 having the formula:


14. A compound according to claim 13 wherein X is C₁-C₆ alkyl; R₂ and R₃are hydrogen; and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which is optionally substituted withone or two R₂₀₀.
 15. A compound according to claim 1 having the formula:


16. A compound according to claim 15 wherein X is C₁-C₆ alkyl; Y isalkynyl, or Y together with the carbon to which it is attached is a D orL amino acid side chain; R₂ and R₃ are hydrogen; and R_(c) is—(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.
 17. A compound according toclaim 1 having the formula:


18. A compound according to claim 17 wherein X is C₁-C₆ alkyl; Y isalkynyl, or Y together with the carbon to which it is attached is a D orL amino acid side chain; R₂ and R₃ are hydrogen; and R_(c) is—(CR₂₄₅R₂₅₀)₀₋₄-aryl or —(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which isoptionally substituted with one or two R₂₀₀.
 19. A compound according toclaim 1 having the formula:


20. A compound according to claim 19 wherein Y is alkynyl, or Y togetherwith the carbon to which it is attached is a D or L amino acid sidechain; R₂ and R₃ are hydrogen; and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which is optionally substituted withone or two R₂₀₀.
 21. A compound according to claim 1 having the formula:


22. A compound according to claim 21 wherein X is C₁-C₆ alkyl; Y isalkynyl, or Y together with the carbon to which it is attached is a D orL amino acid side chain; and R_(c) is —(CR₂₄₅R₂₅₀)₀₋₄-aryl or—(CR₂₄₅R₂₅₀)₀₋₄-heteroaryl, each of which is optionally substituted withone or two R₂₀₀.
 23. A compound according to claim 1 having the formula:


24. A compound according to claim 23 wherein X is C₁-C₆ alkyl; Y isalkynyl, or Y together with the carbon to which it is attached is a D orL amino acid side chain; and R₂₀₀ is C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆alkenyl, trifluoromethyl, or halogen.
 25. A compound according to claim1 having the formula:

wherein V is CH or N.
 26. A compound according to claim 25 wherein X isC₁-C₆ alkyl; Y is alkynyl, or Y together with the carbon to which it isattached is a D or L amino acid side chain; R₂ and R₃ are hydrogen; andR₂₀₀ is C₁-C₆ alkyl, C₂-C₆ alkenyl, C₂-C₆ alkenyl, trifluoromethyl, orhalogen.
 27. A compound according to claim 1 having the formula:


28. A compound according to claim 27 wherein X is C₁-C₆ alkyl; Y isalkynyl, or Y together with the carbon to which it is attached is a D orL amino acid side chain; and R₂ and R₃ are hydrogen.
 29. A compoundaccording to claim 1 having the formula:


30. A compound according to claim 29 wherein X is C₁-C₆ alkyl; Y isalkynyl, or Y together with the carbon to which it is attached is a D orL amino acid side chain; and R₂ and R₃ are hydrogen.
 31. A compoundaccording to claim 1 having the formula:


32. A compound according to claim 31 wherein X is C₁-C₆alkyl; Y isalkynyl, or Y together with the carbon to which it is attached is a D orL amino acid side chain; and R₂ and R₃ are hydrogen.
 33. A compoundaccording to claim 1 which is12-[₂-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1 (17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;2-{12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl}-acetamide;2-(12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-(12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-{16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl}-acetamide;2-(12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-{12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl}-acetamide;2-(12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-(12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-{16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl}-acetamide;2-(12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-{12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl}-acetamide;2-(12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-(12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-{16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl}-acetamide;2-(12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-{12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl}-acetamide;2-(12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-(12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-{16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl}-acetamide;2-(12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-7,10-dioxo-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-trien-9-yl)-acetamide;2-{13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl}-acetamide;2-(13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide;2-(13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide;2-{17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl}-acetamide;2-(13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide;2-{13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl}-acetamide;2-(13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide;2-(13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide;2-{17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl}-acetamide;2-(13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-8,11-dioxo-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-trien-10-yl)-acetamide;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-prop-2-ynyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-10-prop-2-ynyl-2-oxa-9,12-diazabicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-prop-2-ynyl-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-isopropyl-2-oxa-8,11-diazabicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-isopropyl-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.3.1]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;16-Fluoro-12-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;12-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-16-fluoro-8-methyl-9-(2-methylsulfanyl-ethyl)-2-oxa-8,11-diaza-bicyclo[12.2.2]octadeca-1(17),14(18),15-triene-7,10-dione;13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-[2-(3-Ethyl-benzylamino)-1-hydroxy-ethyl]-17-fluoro-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;13-{2-[1-(3-Ethynyl-phenyl)-cyclopropylamino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;17-Fluoro-13-[1-hydroxy-2-(3-trifluoromethyl-benzylamino)-ethyl]-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione;and13-{2-[(5-Ethyl-pyridin-3-ylmethyl)-amino]-1-hydroxy-ethyl}-17-fluoro-9-methyl-10-(2-methylsulfanyl-ethyl)-2-oxa-9,12-diaza-bicyclo[13.3.1]nonadeca-1(18),15(19),16-triene-8,11-dione.